R E S E A R C H T O O L S

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hampton research 34 journey ali so vi ejo, ca 92656- 3317 usa
telephone: 800- 452- 3899 or 949- 425- 1321 fax: 949- 425- 1611
emai l: i nfo@hrmai l. com websi te: www. hamptonresearch. com
v o l u m e 1 8
C R Y S T A L L I Z A T I O N
S O L U T I O N S F O R C R Y S T A L G R O W T H
c o nt a c t i nf o r ma t i o n
Business Hours: 7:00 a.m. to 5:00 p.m.
Monday - Friday
(Pacific Standard Time)
Phone: (800) 452-3899 or
(949) 425-1321

Fax: (949) 425-1611
Address: Hampton Research
34 Journey
Aliso Viejo, CA 92656 U.S.A.
c us t o me r s up p o rt s e rv i c e s
Customer Service: Phone: (800) 452-3899 or
(949) 425-1321
Fax: (949) 425-1611
Email: info@hrmail.com
Technical Support: Phone: (949) 425-1321
Fax: (949) 425-1611
Email: tech@hrmail.com
Website: www.hamptonresearch.com
Solutions for Crystal Growth
On the cover: Crystals of Big-R protein of Xyllela fastidiosa.
Produced in Brazilian Synchrotron Light Laboratory (LNLS),
Center for Structural Molecular Biology by Rosicler Lzaro Barbosa.
T A B L E O F C O N T E N T S
1
On the cover: Crystals of Big-R protein of Xyllela fastidiosa.
Produced in Brazilian Synchrotron Light Laboratory (LNLS),
Center for Structural Molecular Biology by Rosicler Lzaro Barbosa.
2 - 33 s c r e e n s
34 - 37 c u s t o m s h o p c r y s t a l l i z a t i o n r e a g e n t s
38 - 59 o p t i m i z e c r y s t a l l i z a t i o n g r a d e r e a g e n t s
60 - 69 s t o c k o p t i o n s k i t s
70 - 105 c r y s t a l l i z a t i o n p l a t e s , h a r d w a r e & a c c e s s o r i e s
106 -115 t o o l s , s e e d i n g & r e s i n
116 - 135 c r y o c r y s t a l l o g r a p h y
136 - 145 c a p i l l a r y m o u n t s & s u p p l i e s
146 - 153 g o n i o m e t e r h e a d s & s u p p l i e s
154 - 157 x e n o n d e r i v a t i z a t i o n
158 - 163 l a b e l s a n d p e n s
164 - 173 b o o k s
174 - 179 p r o t e i n c r y s t a l l i z a t i o n s t a n d a r d s
180 - 267 c r y s t a l g r o w t h 1 0 1
268 - 275 i n d e x
276 - 277 g e n e r a l i n f o r m a t i o n
P A G E S
Here is a recipe to try:
Mosaicity is about 0.5
t0 0.6
Reagent:
Crystal Screen Cryo
Reagent 23
Mix equal amounts of
Glucose Isomerase
and reagent. Vapor
diffusion method.

Mount crystal in
CryoLoop.

Mosaicity may be
a bit more in this
reagent and the
unit cell will shrink a
s c r e e n s
Empire State Building-shaped crystal of neuron guidance receptor protein grown in Hampton Research Crystal Screen reagent 15.
Momchil Kolev, Dorothea Robev and Dimitar Nikolov, Structural Biology Department at Memorial Sloan-Kettering Cancer Center, New York, USA.
T A B L E O F C O N T E N T S
5 p c t

p r e - c r y s t a l l i z a t i o n t e s t
6 - 7 i n d e x

8 - 9 c r y s t a l s c r e e n

10 - 11 p e g r x

12 - 13 p e g / i o n

14 - 15 g r i d s c r e e n s

q u i k s c r e e n

16 - 17 s a l t r x

18 - 19 m e m b f a c

c r y s t a l s c r e e n l i t e

20 - 21 n a t r i x

22 - 23 c r y s t a l s c r e e n c r y o

24 n u c l e i c a c i d m i n i s c r e e n

25 l o w i o n i c s t r e n g t h s c r e e n

26 - 27 s i l v e r b u l l e t s

28 a d d i t i v e s c r e e n

29 d e t e r g e n t s c r e e n

30 - 32 h e a v y a t o m k i t s

P A G E S
Fish shaped protein crystal grown from
Hampton Research crystallization reagents.
Paul Morin and Kevin Kish, Bristol Myers-Squibb,
Princeton, New Jersey, USA.
Crystals of kinase complex optimized from
a Hampton Research Tacsimate based reagent.
Michelle Quiles, GlaxoSmithKline, RTP,
North Carolina, USA.
Crystals of human mineralocorticoid receptor complexed with ligand.
Grown using Hampton Salt Rx screen.

Kevin P Madauss, GlaxoSmithkline, Research Triangle Park,
North Carolina, USA.
PCT Reagent A1/B1 Results PCT Reagent A2/B2 Results Action
Heavy amorphous precipitate Heavy amorphous precipitate Dilute sample 1:1, repeat steps 1-7
Concentrate sample to half the original
volume, repeat steps 1-7
Clear Clear
Perform screen Light granular precipitate Clear
Perform screen Clear Light granular precipitate
Perform screen Light granular precipitate Light granular precipitate
Heavy amorphous precipitate Clear
Perform PCT with B1 & B2 /
perform diagnostic testing
Clear Heavy amorphous precipitate
Perform PCT with B1 & B2 /
perform diagnostic testing
Figure 1. Light Precipitate
Figure 2. Heavy Amorphous Precipitate
n
Determine the appropriate sample concentra-
tion for crystallization screening
n
Conserve sample
n
Optimize sample concentration prior to initial
screens
n
Provide insight to sample homogeneity
a p p l i c a t i o n
f e a t u r e s
The PCT Pre-Crystallization Test is used
to determine the appropriate sample
concentration for crystallization screen-
ing. Sample concentration is a signifi-
cant crystallization variable. Samples too
concentrated can result in amorphous
precipitate, while samples too dilute
can result in clear drops. Precipitate and
clear drops are typical crystallization screen results for reagent conditions which do not promote crystal-
lization and are part of every crystallization screen. However, by optimizing protein concentration for the
screen, the number of clear and precipitate results can often be reduced, which in turn results in more effi-
cient sample utilization while at the same time enhancing the chances for crystallization. PCT can minimize
or prevent situations where a screen results in an over abundance of precipitate or clear drops.
The PCT kit contains 4 unique, preformulated, sterile filtered reagents used to evaluate protein concentra-
tion for crystallization screening. Initially, the sample protein is mixed with two of the reagents to determine
if the protein concentration is appropriate for crystallization screening. If the protein is very sensitive to salt
and polymer concentration, based on initial PCT results, the protein may be evaluated using a second set
of PCT reagents. PCT results will then provide insight to either the appropriate sample concentration or
indicate that other diagnostic testing such as native gel electrophoresis or dynamic light scattering should
be performed to demonstrate sample homogeneity appropriate for crystallization.
Each PCT kit contains 4 unique reagents, 50 ml each. This kit is useful for researchers who already have
crystallization plates and cover slides.
Each PCT kit (with plates) contains 4 unique reagents, 30 ml each, plus 24 well VDX Plates with sealant
and plain cover slides. This kit is useful for labs without access to crystallization plates and cover slides.
References
1. Crystallization and x-ray diffraction analysis of human CLEC-2. Aleksandra A. Watson and Christopher A. OCallaghan. Acta Cryst. (2005). F61, 10941096
Order Information
Each PCT kit contains 4 unique reagents. To order individual reagents, use Custom Shop

catalog
number listed below. Refer to page 36 for further details.
Cat. No. Name Description Price
HR2-140 PCT 50 ml bottles (4 ea), plain cover slides (1 pk) $80.00
HR2-142 PCT (with Plates) 30 ml bottles (4 ea), plain cover slides (1 pk), $98.00
VDX Plates with sealant (5 ea)
HR2-940-** PCT Custom Shop 185 ml $138.00
** = reagent number A1-B2
d e s c r i p t i o n
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P C T

P R E - C R Y S T A L L I Z A T I O N T E S T
Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com 6
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n
Primary, diverse reagent system crystalliza-
tion screen for proteins, complexes, peptides,
nucleic acids, & water soluble small molecules
n
Developed at Hampton Research
n
A data-driven biased sparse matrix and grid
screen
n
Screens classic, contemporary, & modern
crystallization reagents
n
Samples pH 3 to 9
n
Compatible with microbatch, vapor diffusion,
& liquid diffusion methods
n
Specially formulated reagent zones:
n
Traditional salts versus pH
n
Neutralized organic acids
n
High [salt] with low [polymer]
n
High [polymer] with low [salt]
n
Low ionic strength versus pH
n
PEG & Salt versus pH
n
PEG & Salt
n
Tube or Deep Well block format
a p p l i c a t i o n
f e a t u r e s
Index is designed as a 96 reagent crystallization screen
that combines the strategies of the grid, sparse matrix,
and incomplete factorial screening with traditional, con-
temporary, and new crystallization reagent systems into a
highly effective and efficient format.
Index, as the name implies, efficiently samples a series
of specially formulated reagent zones to identify which reagent class or classes and pH are effective in
producing crystals or limiting sample solubility. Results from Index can be used to design optimization
experiments and to identify follow on screens by reagent class. For example, positive results with salt based
reagent in Index may be followed up with further screening using SaltRx or Grid Screen Salt. Success with
polymer based reagents in Index may be followed up with further screening using PEGRx or PEG/Ion.
Index utilizes a broad, yet refined portfolio of crystallization reagent systems. These include the following:
(1) traditional salts such as Ammonium sulfate and Sodium chloride versus pH; (2) neutralized organic acids
such as Sodium malonate and Tacsimate; (3) High salt concentration mixed with low polymer concentra-
tion as well as high polymer concentration mixed with low salt concentration and; (4) Low ionic strength
using polymers such as PEG, MPD, Pentaerythritols versus pH. These reagent systems are formulated
across a sparse matrix and incomplete factorial of concentration ranges, sampling a pH range of 3 to 9.
Index contains 96 unique reagents, 10 ml each.
Index HT contains 96 unique reagents in a single Deep Well block format.
Ready-to-use reagents are sterile filtered and formulated with ultra-pure Type 1 water, using the highest
purity salts, polymers, organics and buffers.
Measured pH range of kit is 3 to 9 at 25C
Average measured pH of kit is 6.8 at 25C
Median measured pH of kit is 6.9 at 25C
References
1. The advantages of using a modified microbatch method for rapid screening of protein crystallization conditions. A. D'Arcy, A. Mac Sweeney, M. Stihle and
A. Haber. Acta Cryst. (2003). D59, 396-399.
2. Cloning, expression, purification, crystallization and preliminary X-ray diffraction analysis of propionate kinase (TdcD) from Salmonella typhimurium.
D. K. Simanshu and M. R. N. Murthy. Acta Cryst. (2005). F61, 52-55.
3. Preparation, crystallization and preliminary X-ray analysis of the methionine synthase (MetE) from Streptococcus mutans. T.-M. Fu, X.-Y. Zhang, L.-F. Li,
Y.-H. Liang and X.-D. Su. Acta Cryst. (2006). F62, 984-985.
d e s c r i p t i o n
I N D E X

I n d e x H T

Crystals of a diabetes related protein.

Courtesy of Allan DArcy and Aengus Mac Sweeney.
Morphochem AG
s u c c e s s s t o r y
Order Information
Each Index kit contains 96 unique reagents. To order individual reagents, use Custom Shop

cata-
log number listed below. Refer to page 36 for further details.
Cat. No. Name Description Price
HR2-144 Index 10 ml, tube format $570.00
HR2-134 Index HT 1 ml, Deep Well block format $185.00
HR2-944-** Index Custom Shop 185 ml $138.00

** = reagent number 1-96
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1. 0.1 M Citric Acid pH 3.5, 2.0 M Ammonium Sulfate
2. 0.1 M Sodium Acetate trihydrate pH 4.5, 2.0 M Ammonium Sulfate
3. 0.1 M Bis-Tris pH 5.5, 2.0 M Ammonium Sulfate
4. 0.1 M Bis-Tris pH 6.5, 2.0 M Ammonium Sulfate
5. 0.1 M HEPES pH 7.5, 2.0 M Ammonium Sulfate
6. 0.1 M Tris pH 8.5, 2.0 M Ammonium Sulfate
7. 0.1 M Citric Acid pH 3.5, 3.0 M Sodium Chloride
8. 0.1 M Sodium Acetate trihydrate pH 4.5, 3.0 M Sodium Chloride
9. 0.1 M Bis-Tris pH 5.5, 3.0 M Sodium Chloride
10. 0.1 M Bis-Tris pH 6.5, 3.0 M Sodium Chloride
11. 0.1 M HEPES pH 7.5, 3.0 M Sodium Chloride
12. 0.1 M Tris pH 8.5, 3.0 M Sodium Chloride
13. 0.1 M Bis-Tris pH 5.5, 0.3 M Magnesium Formate
14. 0.1 M Bis-Tris pH 6.5, 0.5 M Magnesium Formate
15. 0.1 M HEPES pH 7.5, 0.5 M Magnesium Formate
16. 0.1 M Tris pH 8.5, 0.3 M Magnesium Formate
17. 1.4 M Sodium/Potassium Phosphate pH 5.6
18. 1.4 M Sodium/Potassium Phosphate pH 6.9
19. 1.4 M Sodium/Potassium Phosphate pH 8.2
20. 0.1 M HEPES pH 7.5, 1.4 M tri-Sodium Citrate dihydrate
21. 1.8 M tri-Ammonium Citrate pH 7.0
22. 0.8 M Succinic Acid pH 7.0
23. 2.1 M DL-Malic Acid pH 7.0
24. 2.8 M Sodium Acetate trihydrate pH 7.0
25. 3.5 M Sodium Formate pH 7.0
26. 1.1 M di-Ammonium Tartrate pH 7.0
27. 2.4 M Sodium Malonate pH 7.0
28. 35% v/v Tacsimate pH 7.0
29. 60% v/v Tacsimate pH 7.0
30. 0.1 M Sodium Chloride, 0.1 M Bis-Tris pH 6.5, 1.5 M Ammonium Sulfate
31. 0.8 M Potassium Sodium Tartrate tetrahydrate, 0.1 M Tris pH 8.5, 0.5% w/v Polyethylene Glycol Monomethyl ether 5000
32. 1.0 M Ammonium Sulfate, 0.1 M Bis-Tris pH 5.5, 1% w/v Polyethylene Glycol 3350
33. 1.1 M Sodium Malonate pH 7.0, 0.1 M HEPES pH 7.0, 0.5% v/v Jeffamine ED-2001 Reagent pH 7.0
34. 1.0 M Succinic Acid pH 7.0, 0.1 M HEPES pH 7.0, 1% w/v Polyethylene Glycol Monomethyl ether 2000
35. 1.0 M Ammonium Sulfate, 0.1 M HEPES pH 7.0, 0.5% w/v Polyethylene Glycol 8000
36. 15% v/v Tacsimate pH 7.0, 0.1 M HEPES pH 7.0, 2% w/v Polyethylene Glycol 3350
37. 25% w/v Polyethylene Glycol 1500
38. 0.1 M HEPES pH 7.0, 30% v/v Jeffamine M-600 Reagent pH 7.0
39. 0.1 M HEPES pH 7.0, 30% v/v Jeffamine ED-2001 Reagent pH 7.0
40. 0.1 M Citric Acid pH 3.5, 25% w/v Polyethylene Glycol 3350
41. 0.1 M Sodium Acetate trihydrate pH 4.5, 25% w/v Polyethylene Glycol 3350
42. 0.1 M Bis-Tris pH 5.5, 25% w/v Polyethylene Glycol 3350
43. 0.1 M Bis-Tris pH 6.5, 25% w/v Polyethylene Glycol 3350
44. 0.1 M HEPES pH 7.5, 25% w/v Polyethylene Glycol 3350
45. 0.1 M Tris pH 8.5, 25% w/v Polyethylene Glycol 3350
46. 0.1 M Bis-Tris pH 6.5, 20% w/v Polyethylene Glycol Monomethyl ether 5000
47. 0.1 M Bis-Tris pH 6.5, 28% w/v Polyethylene Glycol Monomethyl ether 2000
48. 0.2 M Calcium Chloride dihydrate, 0.1 M Bis-Tris pH 5.5, 45% v/v 2-Methyl-2,4-pentanediol
49. 0.2 M Calcium Chloride dihydrate, 0.1 M Bis-Tris pH 6.5, 45% v/v 2-Methyl-2,4-pentanediol
50. 0.2 M Ammonium Acetate, 0.1 M Bis-Tris pH 5.5, 45% v/v 2-Methyl-2,4-pentanediol
51. 0.2 M Ammonium Acetate, 0.1 M Bis-Tris pH 6.5, 45% v/v 2-Methyl-2,4-pentanediol
52. 0.2 M Ammonium Acetate, 0.1 M HEPES pH 7.5, 45% v/v 2-Methyl-2,4-pentanediol
53. 0.2 M Ammonium Acetate, 0.1 M Tris pH 8.5, 45% v/v 2-Methyl-2,4-pentanediol
54. 0.05 M Calcium Chloride dihydrate, 0.1 M Bis-Tris pH 6.5, 30% v/v Polyethylene Glycol Monomethyl ether 550
55. 0.05 M Magnesium Chloride hexahydrate, 0.1 M HEPES pH 7.5, 30% v/v Polyethylene Glycol Monomethyl ether 550
56. 0.2 M Potassium Chloride, 0.05 M HEPES pH 7.5, 35% v/v Pentaerythritol Propoxylate (5/4 PO/OH)
57. 0.05 M Ammonium Sulfate, 0.05 M Bis-Tris pH 6.5, 30% v/v Pentaerythritol Ethoxylate (15/4 EO/OH)
58. 0.1 M Bis-Tris pH 6.5, 45% v/v Polypropylene Glycol P 400
59. 0.02 M Magnesium Chloride hexahydrate, 0.1 M HEPES pH 7.5, 22% w/v Polyacrylic Acid 5100 Sodium salt
60. 0.01 M Cobalt Chloride hexahydrate, 0.1 M Tris pH 8.5, 20% w/v Polyvinylpyrrolidone K15
61. 0.2 M Proline, 0.1 M HEPES pH 7.5, 10% w/v Polyethylene Glycol 3350
62. 0.2 M Trimethylamine N-oxide dihydrate, 0.1 M Tris pH 8.5, 20% w/v Polyethylene Glycol Monomethyl ether 2000
63. 5% v/v Tacsimate pH 7.0, 0.1 M HEPES pH 7.0, 10% w/v Polyethylene Glycol Monomethyl ether 5000
64. 0.005 M Cobalt Chloride hexahydrate, 0.005 M Nickel (II) Chloride hexahydrate, 0.005 M Cadmium Chloride dihydrate,
0.005 M Magnesium Chloride hexahydrate, 0.1 M HEPES pH 7.5, 12% w/v Polyethylene Glycol 3350
65. 0.1 M Ammonium Acetate, 0.1 M Bis-Tris pH 5.5, 17% w/v Polyethylene Glycol 10,000
66. 0.2 M Ammonium Sulfate, 0.1 M Bis-Tris pH 5.5, 25% w/v Polyethylene Glycol 3350
67. 0.2 M Ammonium Sulfate, 0.1 M Bis-Tris pH 6.5, 25% w/v Polyethylene Glycol 3350
68. 0.2 M Ammonium Sulfate, 0.1 M HEPES pH 7.5, 25% w/v Polyethylene Glycol 3350
69. 0.2 M Ammonium Sulfate, 0.1 M Tris pH 8.5, 25% w/v Polyethylene Glycol 3350
70. 0.2 M Sodium Chloride, 0.1 M Bis-Tris pH 5.5, 25% w/v Polyethylene Glycol 3350
71. 0.2 M Sodium Chloride, 0.1 M Bis-Tris pH 6.5, 25% w/v Polyethylene Glycol 3350
72. 0.2 M Sodium Chloride, 0.1 M HEPES pH 7.5, 25% w/v Polyethylene Glycol 3350
73. 0.2 M Sodium Chloride, 0.1 M Tris pH 8.5, 25% w/v Polyethylene Glycol 3350
74. 0.2 M Lithium Sulfate monohydrate, 0.1 M Bis-Tris pH 5.5, 25% w/v Polyethylene Glycol 3350
75. 0.2 M Lithium Sulfate monohydrate, 0.1 M Bis-Tris pH 6.5, 25% w/v Polyethylene Glycol 3350
76. 0.2 M Lithium Sulfate monohydrate, 0.1 M HEPES pH 7.5, 25% w/v Polyethylene Glycol 3350
77. 0.2 M Lithium Sulfate monohydrate, 0.1 M Tris pH 8.5, 25% w/v Polyethylene Glycol 3350
78. 0.2 M Ammonium Acetate, 0.1 M Bis-Tris pH 5.5, 25% w/v Polyethylene Glycol 3350
79. 0.2 M Ammonium Acetate, 0.1 M Bis-Tris pH 6.5, 25% w/v Polyethylene Glycol 3350
80. 0.2 M Ammonium Acetate, 0.1 M HEPES pH 7.5, 25% w/v Polyethylene Glycol 3350
81. 0.2 M Ammonium Acetate, 0.1 M Tris pH 8.5, 25% w/v Polyethylene Glycol 3350
82. 0.2 M Magnesium Chloride hexahydrate, 0.1 M Bis-Tris pH 5.5, 25% w/v Polyethylene Glycol 3350
83. 0.2 M Magnesium Chloride hexahydrate, 0.1 M Bis-Tris pH 6.5, 25% w/v Polyethylene Glycol 3350
84. 0.2 M Magnesium Chloride hexahydrate, 0.1 M HEPES pH 7.5, 25% w/v Polyethylene Glycol 3350
85. 0.2 M Magnesium Chloride hexahydrate, 0.1 M Tris HCl pH 8.5, 25% w/v Polyethylene Glycol 3350
86. 0.2 M Potassium Sodium Tartrate tetrahydrate, 20% w/v Polyethylene Glycol 3350
87. 0.2 M Sodium Malonate pH 7.0, 20% w/v Polyethylene Glycol 3350
88. 0.2 M tri-Ammonium Citrate pH 7.0, 20% w/v Polyethylene Glycol 3350
89. 0.1 M Succinic Acid pH 7.0, 15% w/v Polyethylene Glycol 3350
90. 0.2 M Sodium Formate, 20% w/v Polyethylene Glycol 3350
91. 0.15 M DL-Malic Acid pH 7.0, 20% w/v Polyethylene Glycol 3350
92. 0.1 M Magnesium Formate, 15% w/v Polyethylene Glycol 3350
93. 0.05 M Zinc Acetate dihydrate, 20% w/v Polyethylene Glycol 3350
94. 0.2 M tri-Sodium Citrate dihydrate, 20% w/v Polyethylene Glycol 3350
95. 0.1 M Potassium Thiocyanate, 30% w/v Polyethylene Glycol Monomethyl ether 2000
96. 0.15 M Potassium Bromide, 30% w/v Polyethylene Glycol Monomethyl ether 2000
I N D E X F O R M U L A T I O N
A1
A2
A3
A4
A5
A6
A7
A8
A9
A10
A11
A12
B1
B2
B3
B4
B5
B6
B7
B8
B9
B10
B11
B12
C1
C2
C3
C4
C5
C6
C7
C8
C9
C10
C11
C12
D1
D2
D3
D4
D5
D6
D7
D8
D9
D10
D11
D12
E1
E2
E3
E4
E5
E6
E7
E8
E9
E10
E11
E12
F1
F2
F3
F4
F5
F6
F7
F8
F9
F10
F11
F12
G1
G2
G3
G4
G5
G6
G7
G8
G9
G10
G11
G12
H1
H2
H3
H4
H5
H6
H7
H8
H9
H10
H11
H12
A1
A2
A3
A4
A5
A6
A7
A8
A9
A10
A11
A12
B1
B2
B3
B4
B5
B6
B7
B8
B9
B10
B11
B12
C1
C2
C3
C4
C5
C6
C7
C8
C9
C10
C11
C12
D1
D2
D3
D4
D5
D6
D7
D8
D9
D10
D11
D12
E1
E2
E3
E4
E5
E6
E7
E8
E9
E10
E11
E12
F1
F2
F3
F4
F5
F6
F7
F8
F9
F10
F11
F12
G1
G2
G3
G4
G5
G6
G7
G8
G9
G10
G11
G12
H1
H2
H3
H4
H5
H6
H7
H8
H9
H10
H11
H12
Range from 3 to 9
p
H
h i g h [ p o l y m e r ] /
l o w [ s a l t ]
n e u t r a l i z e d
o r g a n i c a c i d s
t r a d i t i o n a l
s a l t s
p o l y m e r /
s a l t
i n d e x
f a c t o r s
h i g h [ s a l t ] /
l o w [ p o l y m e r ]
p o l y m e r / s a l t / p H
l o w i o n i c
s t r e n g t h
n
Primary screen for proteins, soluble peptides,
nucleic acids, & water soluble small molecules
n
Sparse matrix additive screen
n
The original sparse matrix screen
n
Sparse matrix formula efficiently samples
salts, polymers, organics, & pH
n
Proven effective with more than 1,000
biological macromolecules
n
Tube or DeepWell block format
a p p l i c a t i o n
f e a t u r e s
The Crystal Screen and Crystal Screen 2 reagent
kits are designed to provide a highly effective and
rapid screening method for the crystallization
of macromolecules. The screens are simple and
practical for finding initial crystallization condi-
tions. The initial crystallization conditions for
more than 1,000 proteins, peptides, oligonucle-
otides, and small molecules have been deter-
mined using Crystal Screen.
A highly effective approach to overcome the exhaustive search for suitable crystallization conditions is the
use of a sparse matrix method of trial conditions that is biased and selected from known crystallization
conditions for macromolecules. The formulation utilized in Crystal Screen and Crystal Screen 2 evaluates
96 unique mixtures of pH, salts, polymers and organics, and their ability to promote crystal growth.
Crystal Screen contains 50 unique reagents, 10 ml each and is based on the sparse matrix formulation first
described by Jancarik and Kim in 1991.
Crystal Screen 2, an extension of Crystal Screen, contains 48 unique reagents, 10 ml each and is based on
the formulation first described by Cudney et al in 1994.
Crystal Screen HT contains 1 ml each of reagents 1-48 from Crystal Screen and all 48 reagents from Crystal
Screen 2 in a single Deep Well block format.
Ready-to-use reagents are sterile filtered and formulated with ultra-pure Type 1 water, using the highest
purity salts, polymers, organics and buffers.
References
1. Jancarik, J. & Kim, S.H. J. Appl. Cryst. 24, 409-411, (1991).
2. Expression, purification, crystallization and preliminary X-ray analysis of two arginine-biosynthetic enzymes from Mycobacterium tuberculosis.
F. Moradian, C. Garen, L. Cherney, M. Cherney and M. N. G. James. Acta Cryst. (2006). F62, 986-988.
d e s c r i p t i o n
Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com 8
Crystal of gp120, the exterior envelope glycoprotein
of type 1 HIV. Preliminary crystallization conditions
obtained using Crystal Screen from Hampton Research.
Courtesy of P.D. Kwong
1
, R. Wyatt
2
, E. Desjardins
2
,
J. Robinson
3
, F.C. Culp
4
, B.D. Hellmig
4
, R.W. Sweet
4
,
J. Sodroski
2
, and W.A. Hendrickson
1,5
.
1
Columbia University,
2
Dana-Farber Cancer Institute,
3
Tulane
University School of Medicine,
4
GlaxoSmithKline ,
5
HHMI-
Columbia University
s u c c e s s s t o r y
CRYSTAL SCREEN

CrysTal sCreen 2

CrysTal sCreen

HT

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Order Information
Each Crystal Screen kit contains 50 unique reagents. Each Crystal Screen 2 kit contains 48 unique
reagents. To order individual reagents, use Custom Shop

catalog numbers listed below. Refer to

page 36 for further details.
Cat. No. Name Description Price
HR2-110 Crystal Screen 10 ml, tube format $285.00
HR2-112 Crystal Screen 2 10 ml, tube format $285.00
HR2-130 Crystal Screen HT 1 ml, Deep Well block format $185.00
HR2-910-** Crystal Screen Custom Shop 185 ml $138.00
HR2-912-** Crystal Screen 2 Custom Shop 185 ml $138.00

** = reagent number 1-50 (for Crystal Screen)
** = reagent number 1-48 (for Crystal Screen 2)
CRYSTAL SCREEN

CrysTal sCreen 2

CrysTal sCreen

HT

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B1
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B7
B8
B9
B10
B11
B12
C1
C2
C3
C4
C5
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C7
C8
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C11
C12
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H4
H5
H6
H7
H8
H9
H10
H11
H12
C R Y S T A L S C R E E N F O R M U L A T I O N
1. 0.02 M Calcium chloride dihydrate, 0.1 M Sodium acetate trihydrate pH 4.6, 30% v/v (+/-)-2-Methyl-2,4-pentanediol
2. 0.4 M Potassium sodium tartrate tetrahydrate
3. 0.4 M Ammonium phosphate monobasic
4. 0.1 M TRIS hydrochloride pH 8.5, 2.0 M Ammonium sulfate
5. 0.2 M Sodium citrate tribasic dihydrate, 0.1 M HEPES sodium pH 7.5, 30% v/v (+/-)-2-Methyl-2,4-pentanediol
6. 0.2 M Magnesium chloride hexahydrate, 0.1 M TRIS hydrochloride pH 8.5, 30% w/v Polyethylene glycol 4,000
7. 0.1 M Sodium cacodylate trihydrate pH 6.5, 1.4 M Sodium acetate trihydrate
8. 0.2 M Sodium citrate tribasic dihydrate, 0.1 M Sodium cacodylate trihydrate pH 6.5, 30% v/v 2-Propanol
9. 0.2 M Ammonium acetate, 0.1 M Sodium citrate tribasic dihydrate pH 5.6, 30% w/v Polyethylene glycol 4,000
10. 0.2 M Ammonium acetate, 0.1 M Sodium acetate trihydrate pH 4.6, 30% w/v Polyethylene glycol 4,000
11. 0.1 M Sodium citrate tribasic dihydrate pH 5.6, 1.0 M Ammonium phosphate monobasic
12. 0.2 M Magnesium chloride hexahydrate, 0.1 M HEPES sodium pH 7.5, 30% v/v 2-Propanol
13. 0.2 M Sodium citrate tribasic dihydrate, 0.1 M TRIS hydrochloride pH 8.5, 30% v/v Polyethylene glycol 400
14. 0.2 M Calcium chloride dihydrate, 0.1 M HEPES sodium pH 7.5, 28% v/v Polyethylene glycol 400
15. 0.2 M Ammonium sulfate, 0.1 M Sodium cacodylate trihydrate pH 6.5, 30% w/v Polyethylene glycol 8,000
16. 0.1 M HEPES sodium pH 7.5, 1.5 M Lithium sulfate monohydrate
17. 0.2 M Lithium sulfate monohydrate, 0.1 M TRIS hydrochloride pH 8.5, 30% w/v Polyethylene glycol 4,000
18. 0.2 M Magnesium acetate tetrahydrate, 0.1 M Sodium cacodylate trihydrate pH 6.5, 20% w/v Polyethylene glycol 8,000
19. 0.2 M Ammonium acetate, 0.1 M TRIS hydrochloride pH 8.5, 30% v/v 2-Propanol
20. 0.2 M Ammonium sulfate, 0.1 M Sodium acetate trihydrate pH 4.6, 25% w/v Polyethylene glycol 4,000
21. 0.2 M Magnesium acetate tetrahydrate, 0.1 M Sodium cacodylate trihydrate pH 6.5, 30% v/v (+
/
-)-2-Methyl-2,4-pentanediol
22. 0.2 M Sodium acetate trihydrate, 0.1 M TRIS hydrochloride pH 8.5, 30% w/v Polyethylene glycol 4,000
23. 0.2 M Magnesium chloride hexahydrate, 0.1 M HEPES sodium pH 7.5, 30% v/v Polyethylene glycol 400
24. 0.2 M Calcium chloride dihydrate, 0.1 M Sodium acetate trihydrate pH 4.6, 20% v/v 2-Propanol
25. 0.1 M Imidazole pH 6.5, 1.0 M Sodium acetate trihydrate
26. 0.2 M Ammonium acetate, 0.1 M Sodium citrate tribasic dihydrate pH 5.6, 30% v/v (+
/
-)-2-Methyl-2,4-pentanediol
27. 0.2 M Sodium citrate tribasic dihydrate, 0.1 M HEPES sodium pH 7.5, 20% v/v 2-Propanol
28. 0.2 M Sodium acetate trihydrate, 0.1 M Sodium cacodylate trihydrate pH 6.5, 30% w/v Polyethylene glycol 8,000
29. 0.1 M HEPES sodium pH 7.5, 0.8 M Potassium sodium tartrate tetrahydrate
30. 0.2 M Ammonium sulfate, 30% w/v Polyethylene glycol 8,000
31. 0.2 M Ammonium sulfate, 30% w/v Polyethylene glycol 4,000
32. 2.0 M Ammonium sulfate
33. 4.0 M Sodium formate
34. 0.1 M Sodium acetate trihydrate pH 4.6, 2.0 M Sodium formate
35. 0.1 M HEPES sodium pH 7.5, 0.8 M Sodium phosphate monobasic monohydrate, 0.8 M Potassium phosphate monobasic
36. 0.1 M TRIS hydrochloride pH 8.5, 8% w/v Polyethylene glycol 8,000
37. 0.1 M Sodium acetate trihydrate pH 4.6, 8% w/v Polyethylene glycol 4,000
38. 0.1 M HEPES sodium pH 7.5, 1.4 M Sodium citrate tribasic dihydrate
39. 0.1 M HEPES sodium pH 7.5, 2% v/v Polyethylene glycol 400, 2.0 M Ammonium sulfate
40. 0.1 M Sodium citrate tribasic dihydrate pH 5.6, 20% v/v 2-Propanol, 20% w/v Polyethylene glycol 4,000
41. 0.1 M HEPES sodium pH 7.5, 10% v/v 2-Propanol, 20% w/v Polyethylene glycol 4,000
42. 0.05 M Potassium phosphate monobasic, 20% w/v Polyethylene glycol 8,000
43. 30% w/v Polyethylene glycol 1,500
44. 0.2 M Magnesium formate dihydrate
45. 0.2 M Zinc acetate dihydrate, 0.1 M Sodium cacodylate trihydrate pH 6.5, 18% w/v Polyethylene glycol 8,000
46. 0.2 M Calcium acetate hydrate, 0.1 M Sodium cacodylate trihydrate pH 6.5 , 18% w/v Polyethylene glycol 8,000
47. 0.1 M Sodium acetate trihydrate pH 4.6, 2.0 M Ammonium sulfate
48. 0.1 M TRIS hydrochloride pH 8.5, 2.0 M Ammonium phosphate monobasic
49. 1.0 M Lithium sulfate monohydrate, 2% w/v Polyethylene glycol 8,000
50. 0.5 M Lithium sulfate monohydrate, 15% w/v Polyethylene glycol 8,000
1. 2.0 M Sodium chloride, 10% w/v Polyethylene glycol 6,000
2. 0.5 M Sodium chloride, 0.01 M Magnesium chloride hexahydrate, 0.01 M Hexadecyltrimethylammonium bromide
3. 25% v/v Ethylene glycol
4. 35% v/v 1,4-Dioxane
5. 2.0 M Ammonium sulfate, 5% v/v 2-Propanol
6. 1.0 M Imidazole pH 7.0
7. 10% w/v Polyethylene glycol 1,000, 10% w/v Polyethylene glycol 8,000
8. 1.5 M Sodium chloride, 10% v/v Ethanol
9. 0.1 M Sodium acetate trihydrate pH 4.6, 2.0 M Sodium chloride
10. 0.2 M Sodium chloride, 0.1 M Sodium acetate trihydrate pH 4.6, 30% v/v (+
/
-)-2-Methyl-2,4-pentanediol
11. 0.01 M Cobalt(II) chloride hexahydrate, 0.1 M Sodium acetate trihydrate pH 4.6, 1.0 M 1,6-Hexanediol
12. 0.1 M Cadmium chloride hydrate, 0.1 M Sodium acetate trihydrate pH 4.6, 30% v/v Polyethylene glycol 400
13. 0.2 M Ammonium sulfate, 0.1 M Sodium acetate trihydrate pH 4.6, 30% w/v Polyethylene glycol monomethyl ether 2,000
14. 0.2 M Potassium sodium tartrate tetrahydrate, 0.1 M Sodium citrate tribasic dihydrate pH 5.6, 2.0 M Ammonium sulfate
15. 0.5 M Ammonium sulfate, 0.1 M Sodium citrate tribasic dihydrate pH 5.6, 1.0 M Lithium sulfate monohydrate
16. 0.5 M Sodium chloride, 0.1 M Sodium citrate tribasic dihydrate pH 5.6, 2% v/v Ethylene imine polymer
17. 0.1 M Sodium citrate tribasic dihydrate pH 5.6, 35% v/v tert-Butanol
18. 0.01 M Iron(III) chloride hexahydrate, 0.1 M Sodium citrate tribasic dihydrate pH 5.6, 10% v/v Jeffamine M-600
19. 0.1 M Sodium citrate tribasic dihydrate pH 5.6, 2.5 M 1,6-Hexanediol
20. 0.1 M MES monohydrate pH 6.5, 1.6 M Magnesium sulfate heptahydrate
21. 0.1 M Sodium phosphate monobasic monohydrate, 0.1 M Potassium phosphate monobasic,
0.1 M MES monohydrate pH 6.5, 2.0 M Sodium chloride
22. 0.1 M MES monohydrate pH 6.5, 12% w/v Polyethylene glycol 20,000
23. 1.6 M Ammonium sulfate, 0.1 M MES monohydrate pH 6.5, 10% v/v 1,4-Dioxane
24. 0.05 M Cesium chloride, 0.1 M MES monohydrate pH 6.5, 30% v/v Jeffamine M-600
25. 0.01 M Cobalt(II) chloride hexahydrate, 0.1 M MES monohydrate pH 6.5, 1.8 M Ammonium sulfate
26. 0.2 M Ammonium sulfate, 0.1 M MES monohydrate pH 6.5, 30% w/v Polyethylene glycol monomethyl ether 5,000
27. 0.01 M Zinc sulfate heptahydrate, 0.1 M MES monohydrate pH 6.5, 25% v/v Polyethylene glycol monomethyl ether 550
28. 1.6 M Sodium citrate tribasic dihydrate pH 6.5
29. 0.5 M Ammonium sulfate, 0.1 M HEPES pH 7.5, 30% v/v (+
/
-)-2-Methyl-2,4-pentanediol
30. 0.1 M HEPES pH 7.5, 10% w/v Polyethylene glycol 6,000, 5% v/v (+
/
-)-2-Methyl-2,4-pentanediol
31. 0.1 M HEPES pH 7.5, 20% v/v Jeffamine M-600
32. 0.1 M Sodium chloride, 0.1 M HEPES pH 7.5, 1.6 M Ammonium sulfate
33. 0.1 M HEPES pH 7.5, 2.0 M Ammonium formate
34. 0.05 M Cadmium sulfate hydrate, 0.1 M HEPES pH 7.5, 1.0 M Sodium acetate trihydrate
35. 0.1 M HEPES pH 7.5, 70% v/v (+
/
-)-2-Methyl-2,4-pentanediol
36. 0.1 M HEPES pH 7.5, 4.3 M Sodium chloride
37. 0.1 M HEPES pH 7.5, 10% w/v Polyethylene glycol 8,000, 8% v/v Ethylene glycol
38. 0.1 M HEPES pH 7.5, 20% w/v Polyethylene glycol 10,000
39. 0.2 M Magnesium chloride hexahydrate, 0.1 M Tris pH 8.5, 3.4 M 1,6-Hexanediol
40. 0.1 M Tris pH 8.5, 25% v/v tert-Butanol
41. 0.01 M Nickel(II) chloride hexahydrate, 0.1 M Tris pH 8.5, 1.0 M Lithium sulfate monohydrate
42. 1.5 M Ammonium sulfate, 0.1 M Tris pH 8.5, 12% v/v Glycerol
43. 0.2 M Ammonium phosphate monobasic, 0.1 M Tris pH 8.5, 50% v/v (+
/
-)-2-Methyl-2,4-pentanediol
44. 0.1 M Tris pH 8.5, 20% v/v Ethanol
45. 0.01 M Nickel(II) chloride hexahydrate, 0.1 M Tris pH 8.5, 20% w/v Polyethylene glycol monomethyl ether 2,000
46. 0.1 M Sodium chloride, 0.1 M BICINE pH 9.0, 20% v/v Polyethylene glycol monomethyl ether 550
47. 0.1 M BICINE pH 9.0, 2.0 M Magnesium chloride hexahydrate
48. 0.1 M BICINE pH 9.0, 2% v/v 1,4-Dioxane, 10% w/v Polyethylene glycol 20,000
C R Y S T A L S C R E E N 2 F O R M U L A T I O N
Range from 4.0 to 9.0
1,4-Dioxane
Ethanol
2-Propanol
tert-Butanol
1,6-Hexanediol
(+/-)-2-Methyl-2,4-pentanediol
Ethylene glycol
Glycerol
Ethylene imine polymer
Jeffamine
M-600
Polyethylene glycol 400
Polyethylene glycol 1,000
Polyethylene glycol 6,000
Polyethylene glycol 8,000
Polyethylene glycol 10,000
Polyethylene glycol 20,000
Polyethylene glycol MME 550
Polyethylene glycol MME 2,000
Polyethylene glycol MME 5,000
BICINE
HEPES
Imidazole
MES
Sodium acetate
Sodium citrate
TRIS hydrochloride
o r g a n i c
p
H
n o n - v o l a t i l e
o r g a n i c
s a l t
p o l y m e r
b u f f e r
Ammonium phosphate
Potassium phosphate
Sodium phosphate
Ammonium sulfate
Cadmium sulfate
Lithium sulfate
Magnesium sulfate
Zinc sulfate
K/Na tartrate
Hexadecyltrimethylammonium bromide
Sodium acetate
Cadmium chloride
Cesium chloride
Cobalt(II) chloride
Iron(III) chloride
Magnesium chloride
Nickel(II) chloride
Sodium chloride
Sodium citrate
Ammonium formate
p o l y m e r
Range from 4.0 to 9.0
2-Propanol
(+/-)-2-Methyl-2,4-pentanediol
Polyethylene glycol 400
Polyethylene glycol 1,500
Polyethylene glycol 4,000
Polyethylene glycol 8,000
Ammonium acetate
Calcium acetate
Magnesium acetate
Sodium acetate
Zinc acetate
Calcium chloride
Magnesium chloride
Sodium citrate
Magnesium formate
Sodium formate
Ammonium phosphate
Potassium phosphate
Sodium phosphate
Ammonium sulfate
Lithium sulfate
K/Na tartrate
o r g a n i c
p
H
n o n - v o l a t i l e
o r g a n i c
s a l t
HEPES sodium
Imidazole
Sodium acetate
Sodium cacodylate
Sodium citrate
TRIS hydrochloride
b u f f e r
crystal screen
factors
crystal screen 2
factors
n
Primary and secondary, polymer and pH
based crystallization screen for biological
macromolecules
n
Developed at Hampton Research
n
PEGRx 1 primary screen variables are
polymer type (16 different polymers), polymer
molecular weight, pH and low ionic strength.
n
PEGRx 2 primary screen variables are
polymer type (13 different polymers), polymer
molecular weight, pH and secondary reagents
which include additives, salts, volatile
organics and polyols.
n
PEGRx HT combines PEGRx 1 and PEGRx 2
in a single 96 Deep Well block
n
pH range 3.5 - 9, using 10 different buffer
systems
n
Polymer molecular weight range 200 to
20,000
n
Tube or Deep Well block format
a p p l i c a t i o n
f e a t u r e s
PEGRx is a primary and secondary, polymer
and pH based crystallization screen devel-
oped at Hampton Research. It is
designed to evaluate polymer based crystal-
lization reagents and pH in low (PEGRx 1) to
medium ionic strength (PEGRx 2). It is also
designed for use as a secondary or optimiza-
tion screen to follow the Hampton Research
Index screen when polymer based reagents
produce hits and interesting solubility leads.
PEGRx 1 is a crystallization reagent kit designed to evaluate an array of polymers of varying molecu-
lar weight in a low ionic strength environment versus a wide range of pH. Polymer reagents include
Polyethylene glycols, Polyethylene glycol monomethylethers, and Jeffamines. The molecular weight range
between 200 and 20,000 is evaluated in a low ionic strength formulation. Ten different buffers are used to
span the range of pH between 3.5 and 9. The primary screen variables are polymer type, polymer molecular
weight, pH and low ionic strength.
PEGRx 2 is a crystallization reagent kit designed to evaluate an array of polymers of varying molecular
weight in a medium ionic strength environment in the presence of additives, salts, volatile organics and
polyols versus a wide range of pH. Polymer reagents include Polyethylene glycols and Polyethylene gly-
col monomethylethers. The polymer molecular weight range between 200 and 20,000 is evaluated in a
medium ionic strength formulation. Ten different buffers are used to span the range of pH between 3.5 and
9. The primary screen variables are polymer type, polymer molecular weight, pH and secondary reagents
which include additives, salts, volatile organics and polyols.
PEGRx 1 contains 48 unique reagents, 10 ml each.
PEGRx 2 contains 48 unique reagents, 10 ml each.
PEGRx HT contains 1 ml of each reagent from PEGRx 1 and PEGRx 2 in a single Deep Well block format.
Ready-to-use reagents are sterile filtered and formulated with ultra-pure Type 1 water, using the highest
purity salts, polymers, organics and buffers.
d e s c r i p t i o n
Order Information
PEGRx 1 and PEGRx 2 kits each contain 48 unique reagents. To order individual reagents, use
Custom Shop

catalog numbers listed below. Refer to page 36 for further details.

Cat. No. Name Description Price
HR2-082 PEGRx 1 10 ml, tube format $285.00
HR2-084 PEGRx 2 10 ml, tube format $285.00
HR2-086 PEGRx HT 1 ml, Deep Well block format $185.00
HR2-982-** PEGRx 1 Custom Shop 185 ml $138.00
HR2-984-** PEGRx 2 Custom Shop 185 ml $138.00
** = reagent number 1-48
P E G R X 1

P e G r x 2

P e G r x H T

Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com 10
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A2
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A4
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A6
A7
A8
A9
A10
A11
A12
B1
B2
B3
B4
B5
B6
B7
B8
B9
B10
B11
B12
C1
C2
C3
C4
C5
C6
C7
C8
C9
C10
C11
C12
D1
D2
D3
D4
D5
D6
D7
D8
D9
D10
D11
D12
E1
E2
E3
E4
E5
E6
E7
E8
E9
E10
E11
E12
F1
F2
F3
F4
F5
F6
F7
F8
F9
F10
F11
F12
G1
G2
G3
G4
G5
G6
G7
G8
G9
G10
G11
G12
H1
H2
H3
H4
H5
H6
H7
H8
H9
H10
H11
H12
P E G R X 1 F O R M U L A T I O N
1. 0.1 M Citric acid pH 3.5, 34% v/v Polyethylene glycol 200
2. 0.1 M Sodium citrate tribasic dihydrate pH 5.5, 38% v/v Polyethylene glycol 200
3 0.1 M HEPES pH 7.5, 42% v/v Polyethylene glycol 200
4. 0.1 M Sodium acetate trihydrate pH 4.5, 30% v/v Polyethylene glycol 300
5. 0.1 M BIS-TRIS pH 6.5, 25% v/v Polyethylene glycol 300
6. 0.1 M BICINE pH 8.5, 20% v/v Polyethylene glycol 300
7. 0.1 M Sodium acetate trihydrate pH 4.0, 15% v/v Polyethylene glycol 400
8. 0.1 M MES monohydrate pH 6.0, 22% v/v Polyethylene glycol 400
9. 0.1 M Tris pH 8.0, 30% v/v Polyethylene glycol 400
10. 0.1 M Sodium citrate tribasic dihydrate pH 5.0, 30% v/v Polyethylene glycol monomethyl ether 550
11. 0.1 M Imidazole pH 7.0, 25% v/v Polyethylene glycol monomethyl ether 550
12. 0.1 M BIS-TRIS propane pH 9.0, 20% v/v Polyethylene glycol monomethyl ether 550
13. 0.1 M Sodium acetate trihydrate pH 4.0, 10% v/v Jeffamine

M-600

pH 7.0
14. 0.1 M MES monohydrate pH 6.0, 20% v/v Jeffamine

M-600

pH 7.0
15. 0.1 M Tris pH 8.0, 30% v/v Jeffamine

M-600

pH 7.0
16. 0.1 M Citric acid pH 3.5, 14% w/v Polyethylene glycol 1,000
17. 0.1 M Sodium citrate tribasic dihydrate pH 5.5, 22% w/v Polyethylene glycol 1,000
18. 0.1 M HEPES pH 7.5, 30% w/v Polyethylene glycol 1,000
19. 0.1 M Sodium acetate trihydrate pH 4.5, 30% w/v Polyethylene glycol 1,500
20. 0.1 M BIS-TRIS pH 6.5, 20% w/v Polyethylene glycol 1,500
21. 0.1 M BICINE pH 8.5, 15% w/v Polyethylene glycol 1,500
22. 0.1 M Sodium acetate trihydrate pH 4.0, 10% w/v Polyethylene glycol monomethyl ether 2,000
23. 0.1 M MES monohydrate pH 6.0, 20% w/v Polyethylene glycol monomethyl ether 2,000
24. 0.1 M Tris pH 8.0, 30% w/v Polyethylene glycol monomethyl ether 2,000
25. 0.1 M Sodium citrate tribasic dihydrate pH 5.0, 30% v/v Jeffamine

ED-2001 pH 7.0
26. 0.1 M Imidazole pH 7.0, 20% v/v Jeffamine

ED-2001 pH 7.0
27. 0.1 M BIS-TRIS propane pH 9.0, 10% v/v Jeffamine

ED-2001 pH 7.0
28. 0.1 M Citric acid pH 3.5, 25% w/v Polyethylene glycol 3,350
29. 0.1 M Sodium citrate tribasic dihydrate pH 5.5, 18% w/v Polyethylene glycol 3,350
30. 0.1 M HEPES pH 7.0, 12% w/v Polyethylene glycol 3,350
31. 0.1 M Sodium acetate trihydrate pH 4.0, 10% w/v Polyethylene glycol 4,000
32. 0.1 M MES monohydrate pH 6.0, 14% w/v Polyethylene glycol 4,000
33. 0.1 M Tris pH 8.0, 28% w/v Polyethylene glycol 4,000
34. 0.1 M Sodium acetate trihydrate pH 4.5, 30% w/v Polyethylene glycol monomethyl ether 5,000
35. 0.1 M BIS-TRIS pH 6.5, 20% w/v Polyethylene glycol monomethyl ether 5,000
36. 0.1 M BICINE pH 8.5, 8% w/v Polyethylene glycol monomethyl ether 5,000
37. 0.1 M Sodium citrate tribasic dihydrate pH 5.0, 10% w/v Polyethylene glycol 6,000
38. 0.1 M Imidazole pH 7.0, 20% w/v Polyethylene glycol 6,000
39. 0.1 M BIS-TRIS propane pH 9.0, 30% w/v Polyethylene glycol 6,000
40. 0.1 M Citric acid pH 3.5, 28% w/v Polyethylene glycol 8,000
41. 0.1 M Sodium citrate tribasic dihydrate pH 5.5, 16% w/v Polyethylene glycol 8,000
42. 0.1 M HEPES pH 7.5, 4% w/v Polyethylene glycol 8,000
43. 0.1 M Sodium acetate trihydrate pH 4.5, 10% w/v Polyethylene glycol 10,000
44. 0.1 M BIS-TRIS pH 6.5, 16% w/v Polyethylene glycol 10,000
45. 0.1 M BICINE pH 8.5, 20% w/v Polyethylene glycol 10,000
46. 0.1 M Sodium citrate tribasic dihydrate pH 5.0, 18% w/v Polyethylene glycol 20,000
47. 0.1 M Imidazole pH 7.0, 12% w/v Polyethylene glycol 20,000
48. 0.1 M BIS-TRIS propane pH 9.0, 8% w/v Polyethylene glycol 20,000
1. 0.8 M Lithium sulfate monohydrate, 0.1 M Sodium acetate trihydrate pH 4.0, 4% v/v Polyethylene glycol 200
2. 0.2 M Lithium sulfate monohydrate, 0.1 M Sodium citrate tribasic dihydrate pH 5.0, 26% v/v Polyethylene glycol 200
3. 0.05 M Calcium chloride dihydrate, 0.1 M MES monohydrate pH 6.0, 45% v/v Polyethylene glycol 200
4. 28% v/v 2-Propanol, 0.1 M BIS-TRIS pH 6.5, 3% v/v Polyethylene glycol 200
5. 20% v/v Tacsimate pH 7.0, 0.1 M HEPES pH 7.5, 2% v/v Polyethylene glycol 200
6. 10% v/v 2-Propanol, 0.1 M Sodium citrate tribasic dihydrate pH 5.0, 26% v/v Polyethylene glycol 400
7. 0.2 M Ammonium acetate, 0.1 M Sodium citrate tribasic dihydrate pH 5.5, 24% v/v Polyethylene glycol 400
8. 0.2 M Ammonium sulfate, 0.1 M BIS-TRIS pH 6.5, 18% v/v Polyethylene glycol 400
9. 0.19 mM CYMAL

-7, 0.1 M HEPES pH 7.5, 40% v/v Polyethylene glycol 400

10. 6% v/v 2-Propanol, 0.1 M Sodium acetate trihydrate pH 4.5, 26% v/v Polyethylene glycol monomethyl ether 550
11. 1.8 M Ammonium sulfate, 0.1 M BIS-TRIS pH 6.5, 2% v/v Polyethylene glycol monomethyl ether 550
12. 0.15 M DL-Malic acid pH 7.0, 0.1 M Imidazole pH 7.0, 22% v/v Polyethylene glycol monomethyl ether 550
13. 0.1 M Succinic acid pH 7.0, 0.1 M BICINE pH 8.5, 30% v/v Polyethylene glycol monomethyl ether 550
14. 0.1 M Lithium sulfate monohydrate, 0.1 M Sodium citrate tribasic dihydrate pH 5.5, 20% w/v Polyethylene glycol 1,000
15. 0.1 M Sodium malonate pH 8.0, 0.1 M Tris pH 8.0, 30% w/v Polyethylene glycol 1,000
16. 4% v/v (+/-)-2-Methyl-2,4-pentanediol, 0.1 M Citric acid pH 3.5, 20% w/v Polyethylene glycol 1,500
17. 0.2 M L-Proline, 0.1 M HEPES pH 7.5, 24% w/v Polyethylene glycol 1,500
18. 10% v/v 2-Propanol, 0.1 M BICINE pH 8.5, 30% w/v Polyethylene glycol 1,500
19. 0.1 M Sodium chloride, 0.1 M BIS-TRIS propane pH 9.0, 25% w/v Polyethylene glycol 1,500
20. 0.02 M Nickel(II) chloride hexahydrate, 0.02 M Magnesium chloride hexahydrate, 0.02 M Cadmium chloride hydrate,
0.1 M Sodium acetate trihydrate pH 4.5, 24% w/v Polyethylene glycol monomethyl ether 2,000
21. 20% v/v 2-Propanol, 0.1 M MES monohydrate pH 6.0, 20% w/v Polyethylene glycol monomethyl ether 2,000
22. 0.2 M Ammonium citrate tribasic pH 7.0, 0.1 M Imidazole pH 7.0, 20% w/v Polyethylene glycol monomethyl ether 2,000
23. 4.0 M Potassium formate, 0.1 M BIS-TRIS propane pH 9.0, 2% w/v Polyethylene glycol monomethyl ether 2,000
24. 50% v/v Tacsimate pH 4.0, 0.1 M Sodium acetate trihydrate pH 4.5, 1% w/v Polyethylene glycol 3,350
25. 0.10% w/v n-Octyl-b-D-glucoside, 0.1 M Sodium citrate tribasic dihydrate pH 5.5, 22% w/v Polyethylene glycol 3,350
26. 2% v/v Tacsimate pH 7.0, 5% v/v 2-Propanol, 0.1 M Imidazole pH 7.0, 8% w/v Polyethylene glycol 3,350
27. 2% v/v 1,4-Dioxane, 0.1 M Tris pH 8.0, 15% w/v Polyethylene glycol 3,350
28. 18% v/v 2-Propanol, 0.1 M Sodium citrate tribasic dihydrate pH 5.5, 20% w/v Polyethylene glycol 4,000
29. 6% v/v Tacsimate pH 6.0, 0.1 M MES monohydrate pH 6.0, 25% w/v Polyethylene glycol 4,000
30. 0.2 M Magnesium formate dihydrate, 0.1 M Sodium acetate trihydrate pH 4.0,
18% w/v Polyethylene glycol monomethyl ether 5,000
31. 2% v/v Polyethylene glycol 400, 0.1 M Imidazole pH 7.0, 24% w/v Polyethylene glycol monomethyl ether 5,000
32. 0.2 M Sodium formate, 0.1 M BICINE pH 8.5, 20% w/v Polyethylene glycol monomethyl ether 5,000
33. 4% v/v 2-Propanol, 0.1 M BIS-TRIS propane pH 9.0, 20% w/v Polyethylene glycol monomethyl ether 5,000
34. 6% v/v Ethylene glycol, 0.1 M Citric acid pH 3.5, 10% w/v Polyethylene glycol 6,000
35. 0.15 M Lithium sulfate monohydrate, 0.1 M Citric acid pH 3.5, 18% w/v Polyethylene glycol 6,000
36. 10% v/v 2-Propanol, 0.1 M Sodium acetate trihydrate pH 4.0, 22% w/v Polyethylene glycol 6,000
37. 0.2 M Sodium chloride, 0.1 M Sodium acetate trihydrate pH 4.0, 22% w/v Polyethylene glycol 8,000
38. 20% v/v 2-Propanol, 0.1 M Tris pH 8.0, 5% w/v Polyethylene glycol 8,000
39. 10% v/v Polyethylene glycol 200, 0.1 M BIS-TRIS propane pH 9.0, 18% w/v Polyethylene glycol 8,000
40. 15% v/v 2-Propanol, 0.1 M Sodium citrate tribasic dihydrate pH 5.0, 10% w/v Polyethylene glycol 10,000
41. 0.4 M Sodium malonate pH 6.0, 0.1 M MES monohydrate pH 6.0, 0.5% w/v Polyethylene glycol 10,000
42. 0.2 M Potassium sodium tartrate tetrahydrate, 0.1 M BIS-TRIS pH 6.5, 10% w/v Polyethylene glycol 10,000
43. 5% v/v (+/-)-2-Methyl-2,4-pentanediol, 0.1 M HEPES pH 7.5, 10% w/v Polyethylene glycol 10,000
44. 0.2 M Ammonium acetate, 0.1 M Tris pH 8.0, 16% w/v Polyethylene glycol 10,000
45. 5% v/v 2-Propanol, 0.1 M Citric acid pH 3.5, 6% w/v Polyethylene glycol 20,000
46. 1.0 M Sodium malonate pH 5.0, 0.1 M Sodium acetate trihydrate pH 4.5, 2% w/v Polyethylene glycol 20,000
47. 0.2 M Magnesium chloride hexahydrate, 0.1 M Sodium citrate tribasic dihydrate pH 5.0, 10% w/v Polyethylene glycol 20,000
48. 3% w/v Dextran sulfate sodium salt, 0.1 M BICINE pH 8.5, 15% w/v Polyethylene glycol 20,000
P E G R X 2 F O R M U L A T I O N
p o l y m e r
Range from 3.5 to 9
BICINE
BIS-TRIS
BIS-TRIS propane
Citric acid
HEPES
Imidazole
MES monohydrate
Sodium acetate
Sodium citrate
Tris
Jeffamine

ED-2001
Jeffamine

M-600

Polyethylene glycol 200

Polyethylene glycol 300
Polyethylene glycol 400
Polyethylene glycol 1,000
Polyethylene glycol 1,500
Polyethylene glycol 3,350
Polyethylene glycol 4,000
Polyethylene glycol 6,000
Polyethylene glycol 8,000
Polyethylene glycol 10,000
Polyethylene glycol 20,000
Polyethylene glycol MME 550
Polyethylene glycol MME 2,000
Polyethylene glycol MME 5,000
p
H
b u f f e r s
s e c o n d a r y
r e a g e n t s
b u f f e r s
p o l y m e r s
BICINE
BIS-TRIS
BIS-TRIS propane
Citric acid
HEPES
Imidazole
MES monohydrate
Sodium acetate
Sodium citrate
Tris
Range from 3.5 to 9
p
H
Polyethylene glycol 200
Polyethylene glycol 400
Polyethylene glycol 1,000
Polyethylene glycol 1,500
Polyethylene glycol 3,350
Polyethylene glycol 4,000
Polyethylene glycol 6,000
Polyethylene glycol 8,000
Polyethylene glycol 10,000
Polyethylene glycol 20,000
Polyethylene glycol MME 550
Polyethylene glycol MME 2,000
Polyethylene glycol MME 5,000
Additive
Cation
Detergent
Organic Acid
Polyol
Volatile Organic
p e g r x 1
f a c t o r s
p e g r x 2
f a c t o r s
n
Primary or secondary, polymer, salt and pH
matrix crystallization screen for biological
macromolecules
n
Developed at Hampton Research
n
PEG/Ion is a sparse matrix profile of anions
and cations in the presence of monodisperse
Polyethylene glycol 3,350 over pH 4.5 - 9.2
n
PEG/Ion 2 screens a complete profile of
titrated organic acids at varying pH levels
(3.7 - 8.8) in the presence of monodisperse
PEG 3,350
n
PEG/Ion HT combines PEG/Ion and PEG/Ion
2 in a single 96 Deep Well block
n
Tube or Deep Well block format
a p p l i c a t i o n
f e a t u r e s
PEG/Ion, developed by Hampton Research,
is a crystallization screen designed to evalu-
ate monodisperse, high purity Polyethylene
glycol 3,350 and 48 unique salts represent-
ing a very complete range of anions and
cations frequently used in the crystallization
of biological macromolecules. The primary
screening variables are PEG, ion type, ionic
strength, and pH. More than 60% of the
published crystallizations utilized PEG as a
primary crystallization reagent and in approx-
imately 50% of those reports, the PEG was combined with an ion as a secondary crystallization reagent.
PEG/Ion 2 is an extension to the fundamental crystallization strategy in PEG/Ion. PEG/Ion 2 reagents cover
the monodisperse, high purity Polyethylene glycol 3,350 and an array of neutralized and pH adjusted
organic acids, multivalent ions, a novel Citrate BIS-TRIS propane buffer system and pH (4 - 8.8). The
formulation of PEG/Ion 2 was developed at Hampton Research. Each of the 48 reagents in PEG/Ion 2
contains PEG 3,350 as the polymer (precipitant). The concentration of PEG is varied from 12% w/v to 20%
w/v depending upon the type and concentration of buffer/salt paired with the polymer. Thirteen of the
forty-eight PEG/Ion 2 reagents contain a separate buffer component. The remaining PEG/Ion 2 reagents
are buffered by the titrated organic acid salt. Six of these thirteen conditions feature a novel Citric acid BIS-
TRIS propane (CBTP) buffer. The CBTP buffer uses Citric acid and BIS-TRIS propane as the acid base pair
to create a two component buffer system effective across pH 2.5 to 9.5. The ratio of Citric acid to BIS-TRIS
propane determines the solution pH. Thirty-five of the forty-eight PEG/Ion 2 reagents contain a neutralized
or pH adjusted organic acid in the presence of the polymer. Neutralized organic acids are highly effective
crystallization salts.
1
Four PEG/Ion 2 reagents feature polyvalent cations. Two of these reagents contain
cation mixes, saving sample by screening six different cations with only two reagents. Tryptone, a casein
digest combinatorial library of peptides, is included in PEG/Ion 2.
PEG/Ion contains 48 unique reagents, 10 ml each.
PEG/Ion 2 contains 48 unique reagents, 10 ml each.
PEG/Ion HT contains 1 ml of each reagent from PEG/Ion and PEG/Ion 2 in a single Deep Well block
format.
Ready-to-use reagents are sterile filtered and formulated with ultra-pure Type 1 water, using the highest
purity salts, polymers, organics and buffers.
PEG/Ion 2
Measured pH range of kit is 3.7 to 8.8 at 25C
Average measured pH of kit is 6.4 at 25C
Median measured pH of kit is 6.7 at 25C
Mode measured pH of kit is 6.7 at 25C
References
1. A comparison of salts for the crystallization of macromolecules. Alexander McPherson. Protein Science (2001), 10:418-422.
2. Searching for silver bullets: An alternative strategy for crystallizing macromolecules. Alexander McPherson and Bob Cudney. Journal of Structural Biology
Volume 156, Issue 3 , December 2006, Pages 387-406
d e s c r i p t i o n
Order Information
PEG/Ion Screen and PEG/Ion 2 Screen kits each contain 48 unique reagents. To order individual
reagents, use Custom Shop

catalog numbers listed below. Refer to page 36 for further details.

Cat. No. Name Description Price
HR2-126 PEG/Ion Screen 10 ml, tube format $285.00
HR2-098 PEG/Ion 2 Screen 10 ml, tube format $285.00
HR2-139 PEG/Ion HT 1 ml, Deep Well block format $185.00
HR2-922-** PEG/Ion Screen Custom Shop 185 ml $138.00
HR2-998-** PEG/Ion 2 Screen Custom Shop 185 ml $138.00
** = reagent number 1-48
Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com 12
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Crystals of Proteinase K. Preliminary crystallization con-
ditions obtained using PEG/Ion Screen from Hampton
Research.
Crystals grown at Hampton Research.
s u c c e s s s t o r y
P E G / I O N S C R E E N

P e G / I o n 2 s C r e e n

P e G / I o n H T

13
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A1
A2
A3
A4
A5
A6
A7
A8
A9
A10
A11
A12
B1
B2
B3
B4
B5
B6
B7
B8
B9
B10
B11
B12
C1
C2
C3
C4
C5
C6
C7
C8
C9
C10
C11
C12
D1
D2
D3
D4
D5
D6
D7
D8
D9
D10
D11
D12
E1
E2
E3
E4
E5
E6
E7
E8
E9
E10
E11
E12
F1
F2
F3
F4
F5
F6
F7
F8
F9
F10
F11
F12
G1
G2
G3
G4
G5
G6
G7
G8
G9
G10
G11
G12
H1
H2
H3
H4
H5
H6
H7
H8
H9
H10
H11
H12
P E G / I O N S C R E E N F O R M U L A T I O N
1. 0.2 M Sodium uoride, 20% w/v Polyethylene glycol 3,350
2. 0.2 M Potassium uoride, 20% w/v Polyethylene glycol 3,350
3. 0.2 M Ammonium uoride, 20% w/v Polyethylene glycol 3,350
4. 0.2 M Lithium chloride, 20% w/v Polyethylene glycol 3,350
5. 0.2 M Magnesium chloride hexahydrate, 20% w/v Polyethylene glycol 3,350
6. 0.2 M Sodium chloride, 20% w/v Polyethylene glycol 3,350
7. 0.2 M Calcium chloride dihydrate, 20% w/v Polyethylene glycol 3,350
8. 0.2 M Potassium chloride, 20% w/v Polyethylene glycol 3,350
9. 0.2 M Ammonium chloride, 20% w/v Polyethylene glycol 3,350
10. 0.2 M Sodium iodide, 20% w/v Polyethylene glycol 3,350
11. 0.2 M Potassium iodide, 20% w/v Polyethylene glycol 3,350
12. 0.2 M Ammonium iodide, 20% w/v Polyethylene glycol 3,350
13. 0.2 M Sodium thiocyanate, 20% w/v Polyethylene glycol 3,350
14. 0.2 M Potassium thiocyanate, 20% w/v Polyethylene glycol 3,350
15. 0.2 M Lithium nitrate, 20% w/v Polyethylene glycol 3,350
16. 0.2 M Magnesium nitrate hexahydrate, 20% w/v Polyethylene glycol 3,350
17. 0.2 M Sodium nitrate, 20% w/v Polyethylene glycol 3,350
18. 0.2 M Potassium nitrate, 20% w/v Polyethylene glycol 3,350
19. 0.2 M Ammonium nitrate, 20% w/v Polyethylene glycol 3,350
20. 0.2 M Magnesium formate dihydrate, 20% w/v Polyethylene glycol 3,350
21. 0.2 M Sodium formate, 20% w/v Polyethylene glycol 3,350
22. 0.2 M Potassium formate, 20% w/v Polyethylene glycol 3,350
23. 0.2 M Ammonium formate, 20% w/v Polyethylene glycol 3,350
24. 0.2 M Lithium acetate dihydrate, 20% w/v Polyethylene glycol 3,350
25. 0.2 M Magnesium acetate tetrahydrate, 20% w/v Polyethylene glycol 3,350
26. 0.2 M Zinc acetate dihydrate, 20% w/v Polyethylene glycol 3,350
27. 0.2 M Sodium acetate trihydrate, 20% w/v Polyethylene glycol 3,350
28. 0.2 M Calcium acetate hydrate, 20% w/v Polyethylene glycol 3,350
29. 0.2 M Potassium acetate, 20% w/v Polyethylene glycol 3,350
30. 0.2 M Ammonium acetate, 20% w/v Polyethylene glycol 3,350
31. 0.2 M Lithium sulfate monohydrate, 20% w/v Polyethylene glycol 3,350
32. 0.2 M Magnesium sulfate heptahydrate, 20% w/v Polyethylene glycol 3,350
33. 0.2 M Sodium sulfate decahydrate, 20% w/v Polyethylene glycol 3,350
34. 0.2 M Potassium sulfate, 20% w/v Polyethylene glycol 3,350
35. 0.2 M Ammonium sulfate, 20% w/v Polyethylene glycol 3,350
36. 0.2 M Sodium tartrate dibasic dihydrate, 20% w/v Polyethylene glycol 3,350
37. 0.2 M Potassium sodium tartrate tetrahydrate, 20% w/v Polyethylene glycol 3,350
38. 0.2 M Ammonium tartrate dibasic, 20% w/v Polyethylene glycol 3,350
39. 0.2 M Sodium phosphate monobasic monohydrate, 20% w/v Polyethylene glycol 3,350
40. 0.2 M Sodium phosphate dibasic dihydrate, 20% w/v Polyethylene glycol 3,350
41. 0.2 M Potassium phosphate monobasic, 20% w/v Polyethylene glycol 3,350
42. 0.2 M Potassium phosphate dibasic, 20% w/v Polyethylene glycol 3,350
43. 0.2 M Ammonium phosphate monobasic, 20% w/v Polyethylene glycol 3,350
44. 0.2 M Ammonium phosphate dibasic, 20% w/v Polyethylene glycol 3,350
45. 0.2 M Lithium citrate tribasic tetrahydrate, 20% w/v Polyethylene glycol 3,350
46. 0.2 M Sodium citrate tribasic dihydrate, 20% w/v Polyethylene glycol 3,350
47. 0.2 M Potassium citrate tribasic monohydrate, 20% w/v Polyethylene glycol 3,350
48. 0.2 M Ammonium citrate dibasic, 20% w/v Polyethylene glycol 3,350

1. 0.1 M Sodium malonate pH 4.0, 12% w/v Polyethylene glycol 3,350
2. 0.2 M Sodium malonate pH 4.0, 20% w/v Polyethylene glycol 3,350
3. 0.1 M Sodium malonate pH 5.0, 12% w/v Polyethylene glycol 3,350
4. 0.2 M Sodium malonate pH 5.0, 20% w/v Polyethylene glycol 3,350
5. 0.1 M Sodium malonate pH 6.0, 12% w/v Polyethylene glycol 3,350
6. 0.2 M Sodium malonate pH 6.0, 20% w/v Polyethylene glycol 3,350
7. 0.1 M Sodium malonate pH 7.0, 12% w/v Polyethylene glycol 3,350
8. 0.2 M Sodium malonate pH 7.0, 20% w/v Polyethylene glycol 3,350
9. 4% v/v Tacsimate pH 4.0, 12% w/v Polyethylene glycol 3,350
10. 8% v/v Tacsimate pH 4.0, 20% w/v Polyethylene glycol 3,350
11. 4% v/v Tacsimate pH 5.0, 12% w/v Polyethylene glycol 3,350
12. 8% v/v Tacsimate pH 5.0, 20% w/v Polyethylene glycol 3,350
13. 4% v/v Tacsimate pH 6.0, 12% w/v Polyethylene glycol 3,350
14. 8% v/v Tacsimate pH 6.0, 20% w/v Polyethylene glycol 3,350
15. 4% v/v Tacsimate pH 7.0, 12% w/v Polyethylene glycol 3,350
16. 8% v/v Tacsimate pH 7.0, 20% w/v Polyethylene glycol 3,350
17. 4% v/v Tacsimate pH 8.0, 12% w/v Polyethylene glycol 3,350
18. 8% v/v Tacsimate pH 8.0, 20% w/v Polyethylene glycol 3,350
19. 0.1 M Succinic acid pH 7.0, 12% w/v Polyethylene glycol 3,350
20. 0.2 M Succinic acid pH 7.0, 20% w/v Polyethylene glycol 3,350
21. 0.1 M Ammonium citrate tribasic pH 7.0, 12% w/v Polyethylene glycol 3,350
22. 0.2 M Ammonium citrate tribasic pH 7.0, 20% w/v Polyethylene glycol 3,350
23. 0.1 M DL-Malic acid pH 7.0, 12% w/v Polyethylene glycol 3,350
24. 0.2 M DL-Malic acid pH 7.0, 20% w/v Polyethylene glycol 3,350
25. 0.1 M Sodium acetate trihydrate pH 7.0, 12% w/v Polyethylene glycol 3,350
26. 0.2 M Sodium acetate trihydrate pH 7.0, 20% w/v Polyethylene glycol 3,350
27. 0.1 M Sodium formate pH 7.0, 12% w/v Polyethylene glycol 3,350
28. 0.2 M Sodium formate pH 7.0, 20% w/v Polyethylene glycol 3,350
29. 0.1 M Ammonium tartrate dibasic pH 7.0, 12% w/v Polyethylene glycol 3,350
30. 0.2 M Ammonium tartrate dibasic pH 7.0, 20% w/v Polyethylene glycol 3,350
31. 2% v/v Tacsimate pH 4.0, 0.1 M Sodium acetate trihydrate pH 4.6, 16% w/v Polyethylene glycol 3,350
32. 2% v/v Tacsimate pH 5.0, 0.1 M Sodium citrate tribasic dihydrate pH 5.6, 16% w/v Polyethylene glycol 3,350
33. 2% v/v Tacsimate pH 6.0, 0.1 M BIS-TRIS pH 6.5, 20% w/v Polyethylene glycol 3,350
34. 2% v/v Tacsimate pH 7.0, 0.1 M HEPES pH 7.5, 20% w/v Polyethylene glycol 3,350
35. 2% v/v Tacsimate pH 8.0, 0.1 M Tris pH 8.5, 16% w/v Polyethylene glycol 3,350
36. (0.07 M Citric acid, 0.03 M BIS-TRIS propane / pH 3.4), 16% w/v Polyethylene glycol 3,350
37. (0.06 M Citric acid, 0.04 M BIS-TRIS propane / pH 4.1), 16% w/v Polyethylene glycol 3,350
38. (0.05 M Citric acid, 0.05 M BIS-TRIS propane / pH 5.0), 16% w/v Polyethylene glycol 3,350
39. (0.04 M Citric acid, 0.06 M BIS-TRIS propane / pH 6.4), 20% w/v Polyethylene glycol 3,350
40. (0.03 M Citric acid, 0.07 M BIS-TRIS propane / pH 7.6), 20% w/v Polyethylene glycol 3,350
41. (0.02 M Citric acid, 0.08 M BIS-TRIS propane / pH 8.8), 16% w/v Polyethylene glycol 3,350
42. 0.02 M Calcium chloride dihydrate, 0.02 M Cadmium chloride hydrate,
0.02 M Cobalt(II) chloride hexahydrate, 20% w/v Polyethylene glycol 3,350
43. 0.01 M Magnesium chloride hexahydrate, 0.005 M Nickel(II) chloride hexahydrate
0.1 M HEPES sodium pH 7.0, 15% w/v Polyethylene glycol 3,350
44. 0.02 M Zinc chloride, 20% w/v Polyethylene glycol 3,350
45. 0.15 M Cesium chloride, 15% w/v Polyethylene glycol 3,350
46. 0.2 M Sodium bromide, 20% w/v Polyethylene glycol 3,350
47. 1% w/v Tryptone, 0.05 M HEPES sodium pH 7.0, 12% w/v Polyethylene glycol 3,350
48. 1% w/v Tryptone, 0.05 M HEPES sodium pH 7.0, 20% w/v Polyethylene glycol 3,350
P E G / I O N 2 S C R E E N F O R M U L A T I O N
m u l t i v a l e n t
i o n s
b u f f e r s
o r g a n i c s a l t s
Neutralized Organic Salts
BIS-TRIS
Citric acid BIS-TRIS propane (CBTP)
HEPES
HEPES sodium
Sodium acetate
Sodium citrate
Tris
Range from 4 to 8
p
H
Ammonium tartrate
Ammonium citrate
Malic acid
Succinic acid
Sodium acetate
Sodium formate
Sodium malonate
Tacsimate

Range from 4.0 to 9.0

Polyethylene glycol 3,350
Acetate
Citrate
Chloride
Fluoride
Formate
Iodide
Nitrate
Phosphate
Sulfate
Thiocyanate
Tartrate
p
H
a n i o n
p o l y m e r
Ammonium
Calcium
Lithium
Magnesium
Potassium
Sodium
Zinc
c a t i o n
Polyethylene glycol 3,350
p o l y m e r
Cadmium chloride
Calcium chloride
Cesium chloride
Cobalt(II) chloride
Magnesium chloride
Nickel(II) chloride
Sodium bromide
Zinc chloride
p e p t i d e s /
a m i n o a c i d s
Tryptone
p e g / i o n
f a c t o r s
p e g / i o n 2
f a c t o r s
Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com 14
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Grid Screen Salt HT combines Grid Screen
Ammonium Sulfate, Grid Screen Sodium
Malonate, Quik Screen and Grid Screen
Sodium Chloride into a single 96 Deep Well
block format. The block contains 1 ml of
each reagent.
Grid Screen Ammonium Sulfate systemati-
cally evaluates Ammonium sulfate at four
concentrations (0.8, 1.6, 2.4, 3.2 M) versus 6
pH levels (4, 5, 6, 7, 8, 9).
Grid Screen Sodium Malonate evaluates Sodium malonate at six concentrations (1.0, 1.5, 1.9, 2.4, 2.9,
3.4 M) versus four pH levels (4, 5, 6, 7).
Quik Screen evaluates Sodium potassium phosphate at four concentrations (0.8, 1.0, 1.4, 1.8 M) versus
six pH levels (5.0, 5.6, 6.3, 6.9, 7.5, 8.2).
Grid Screen Sodium Chloride evaluates Sodium chloride at four concentrations (1.0, 2.0, 3.0, 4.0 M)
versus six pH levels (4, 5, 6, 7, 8, 9).
Grid Screen PEG 6000 evaluates Polyethylene glycol at four concentrations (5, 10, 20, 30 %w/v) versus
six pH levels (4, 5, 6, 7, 8, 9).
Grid Screen MPD evaluates MPD at four concentrations (10, 20, 40, 65 %v/v) versus six pH levels (4,
5, 6, 7, 8, 9).
Grid Screen PEG/LiCl evaluates Polyethylene glycol at four concentrations (0, 10, 20, 30 %w/v) in the
presence of 1.0 M Lithium chloride versus six pH levels (4, 5, 6, 7, 8, 9).
Grid Screens and Quik Screen each contain 10 ml of 24 unique reagents.
Ready-to-use reagents are sterile filtered and formulated with ultra-pure Type 1 water, using the high-
est purity salts, polymers, organics and buffers.
References
1. A protein crystallization strategy using automated grid searches on successively finer grids. Patricia C. Weber. Methods: A Companion to Methods in
Enzymology Vol. 1, No. 1, August, pp. 31-37, 1990.
2. Protein Crystallization; Techniques, Strategies, and Tips. A Laboratory Manual. Edited by Terese M. Bergfors. International University Line, 1999. ISBN
0-9636817-5-3.
d e s c r i p t i o n
a p p l i c a t i o n
f e a t u r e s
n
Primary or secondary, salt, polymer, organic
and pH grid crystallization screen for biologi-
cal macromolecules
n
A systematic grid screen varying salt or poly-
mer or organic versus pH
n
Samples pH 4 to 9
n
Combine reagents within or between screens
to customize your screen
n
Combine reagents between rows or columns
to create expanded grid screens
n
Tube or Deep Well block format
Order Information
Each Grid Screen Salt HT kit contains 96 unique reagents.
Cat. No. Name Description Price
HR2-248 Grid Screen Salt HT 1 ml, Deep Well block format $185.00
G R I D S C R E E N S

q u I k s C r e e n

Number of crystals grown
versus varying A/S
concentrations [M]
A
m
m
oni um
S
ul f at e [ M
]
60
50
40
30
20
10
0
0
10
20
30
40
50
60
0
to
5
6
to
1
0
1
1
to
1
6
1
6
to
2
0
2
1
to
2
5
2
5
to
3
0
3
0
+
C
ry
s
ta
ls
G
ro
w
n
N
u
m
b
e
r

o
f

C
r
y
s
t
a
l
s
%
w
/v P
E
G
6000
Number of crystals grown
versus varying PEG 6000
concentrations (% w/v)
7
0

t
o

7
9
6
0

t
o

6
9
5
0

t
o

5
9
4
0

t
o

4
9
3
0

t
o

3
9
2
0

t
o

2
9
1
0

t
o

1
9
0

t
o

9
C
rystals G
ro
w
n
30
25
20
15
10
5
0
N
u
m
b
e
r

o
f

C
r
y
s
t
a
l
s
%
v/v M
PD
0
5
10
15
20
25
30
Number of crystals grown
versus varying MPD
concentrations (% v/v)
15
s
c
r
e
e
n
s
Cat. No. Name Description Price
HR2-211 Grid Screen Ammonium Sulfate 10 ml, tube format $175.00
HR2-924-** Grid Screen Ammonium Sulfate Custom Shop 185 ml $138.00
** = reagent number A1-D6
Order Information
Each Grid Screen and Quik Screen kit contains 24 unique reagents. To order individual reagents, use Custom Shop

catalog number
listed below. Refer to page 36 for further details.
Cat. No. Name Description Price
HR2-247 Grid Screen Sodium Malonate 10 ml, tube format $175.00
HR2-947-** Grid Screen Sodium Malonate Custom Shop 185 ml $138.00
** = reagent number A1-D6
Cat. No. Name Description Price
HR2-221 Quik Screen 10 ml, tube format $175.00
HR2-921-** Quik Screen Custom Shop 185 ml $138.00
** = reagent number A1-D6
Cat. No. Name Description Price
HR2-219 Grid Screen Sodium Chloride 10 ml, tube format $175.00
HR2-932-** Grid Screen Sodium Chloride Custom Shop 185 ml $138.00
** = reagent number A1-D6
Cat. No. Name Description Price
HR2-213 Grid Screen PEG 6000 10 ml, tube format $175.00
HR2-926-** Grid Screen PEG 6000 Custom Shop 185 ml $138.00
** = reagent number A1-D6
Cat. No. Name Description Price
HR2-215 Grid Screen MPD 10 ml, tube format $175.00
HR2-930-** Grid Screen MPD Custom Shop 185 ml $138.00
** = reagent number A1-D6
Cat. No. Name Description Price
HR2-217 Grid Screen PEG/LiCl 10 ml, tube format $175.00
HR2-928-** Grid Screen PEG/LiCl Custom Shop 185 ml $138.00
** = reagent number A1-D6
[ M ]
Sodium malonate
1
1.0
2
1.5
3
1.9
4
2.4
5
2.9
6
3.4
B
A
C
5.0
4.0
6.0
7.0 D
pH
[ M ]
Sodium/Potassium
Phosphate
1
5.0
2
5.6
3
6.3
4
6.9
5
7.5
6
8.2
B
A
C
1.0 M
0.8 M
1.4 M
1.8 M D
pH
0.1 M Buffer
[ M ]
Ammonium
sulfate
1
4
2
5
3
6
4
7
5
8
6
9
1.6 M B
0.8 M A
2.4 M C
3.0 M D
C
itr
ic
a
c
id
M
E
S
H
E
P
E
S
T
r
is
B
IC
IN
E
C
itr
ic
a
c
id
[ % w/v ]
Polyethylene
glycol
6,000
0.1 M Buffer
1
4
2
5
3
6
4
7
5
8
6
9
10% B
5% A
20% C
30% D
C
itr
ic
a
c
id
M
E
S
H
E
P
E
S
T
r
is
B
IC
IN
E
C
itr
ic
a
c
id
[ M ]
Sodium
chloride
0.1 M Buffer
1
4
2
5
3
6
4
7
5
8
6
9
2.0 M B
1.0 M A
3.0 M C
4.0 M D
C
itr
ic
a
c
id
M
E
S
H
E
P
E
S
T
r
is
B
IC
IN
E
C
itr
ic
a
c
id
[ % v/v ]
MPD
0.1 M Buffer
1
4
2
5
3
6
4
7
5
8
6
9
20% B
10% A
40% C
65% D
C
itr
ic
a
c
id
M
E
S
H
E
P
E
S
T
r
is
B
IC
IN
E
C
itr
ic
a
c
id
[ % w/v ]
Polyethylene
glycol
6,000
1.0 M Lithium chloride in all reagents
0.1 M Buffer
1
4
2
5
3
6
4
7
5
8
6
9
10% B
5% A
20% C
30% D
C
itr
ic
a
c
id
M
E
S
H
E
P
E
S
T
r
is
B
IC
IN
E
C
itr
ic
a
c
id
Crystals grown using SaltRx, reagent 62. They were
grown using the Cyberlab C240 robot and the Neuro
Probe plate. These were grown just in time to freeze
and take to the synchrotron (no time for optimizations).
MAD data were collected on these crystals and the
structure has been solved to 2.3 resolution.
Courtesy of Annie Hassell.
GlaxoSmithKline
s u c c e s s s t o r y
n
Primary or secondary, salt and pH matrix
crystallization screen for biological macromol-
ecules
n
Developed at Hampton Research
n
Salt versus pH matrix crystallization screen
n
Samples pH 4 - 9
n
22 unique salts versus salt concentration
and pH
n
Compatible with microbatch, vapor diffusion,
liquid & gel diffusion methods
n
Preformulated, ready to screen
n
All salts and buffers in screen readily avail-
able as Optimize reagents for reproducing
and optimizing crystals
n
Tube or Deep Well block format
a p p l i c a t i o n
f e a t u r e s
SaltRx was developed by Hampton Research
as a primary and secondary, salt and pH
based crystallization screen for biological
macromolecules. Salt is the only primary
crystallization reagent (precipitant) utilized.
Based on a design of 96 conditions, the
screen evaluates a broad portfolio of crystalli-
zation salts of varying concentration and pH.
The selection, concentration, and pH of the
salts were determined by data mining the BMCD
1
and other crystallization databases, crystallization reports
in the literature, and internal crystallization trials performed at Hampton Research. Based on this analysis,
up to 35% of protein crystallizations involve salt as the primary crystallization reagent.
SaltRx can be used as a primary crystallization screen when salt, ionic strength and pH are desired or
suspected as appropriate crystallization variables. It is also useful as a secondary screen when salt based
reagents/conditions from screens such as Index, Crystal Screen, and Grid Screen produce crystals and
when further screening for additional salt conditions or optimization is desired.
SaltRx is designed to be used as a 96 reagent screen.
SaltRx 1 contains 48 unique reagents, 10 ml each.
SaltRx 2 contains 48 unique reagents, 10 ml each.
SaltRx HT contains 1 ml of each reagent from SaltRx 1 and SaltRx 2 in a single Deep Well block format.
Ready-to-use reagents are sterile filtered and formulated with ultra-pure Type 1 water, using the highest
purity salts, polymers, organics and buffers.
References
1. Gilliland, G.L., Tung, M., Blakeslee, D.M. and Ladner, J. 1994. The Biological Macromolecule Crystallization Database, Version 3.0: New
Features, Data, and the NASAArchive for Protein Crystal Growth Data. Acta Crystallogr. D50 408-413.
2. Crystallization and preliminary crystallographic analysis of molybdenumcofactor biosynthesis protein C from Thermus thermophilus. S. P. Kanaujia, C. V.
Ranjani, J. Jeyakanthan, S. Baba, L. Chen, Z.-J. Liu, B.-C. Wang, M. Nishida, A. Ebihara, A. Shinkai, S. Kuramitsu, Y. Shiro,
K. Sekar and S. Yokoyama. Acta Cryst. (2007). F63, 27-29.
Order Information
SaltRx 1 and SaltRx 2 kits each contain 48 unique reagents. To order individual reagents, use
Custom Shop

catalog number listed below. Refer to page 36 for further details.

Cat. No. Name Description Price
HR2-107 SaltRx 1 10 ml, tube format $285.00
HR2-109 SaltRx 2 10 ml, tube format $285.00
HR2-136 SaltRx HT 1 ml, Deep Well block format $185.00
HR2-907-** SaltRx 1 Custom Shop 185 ml $138.00
HR2-909-** SaltRx 2 Custom Shop 185 ml $138.00
** = reagent number 1-48
d e s c r i p t i o n
Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com 16
s
c
r
e
e
n
s
S A L T R X 1

s a l T r x 2

s a l T r x H T

17
s
c
r
e
e
n
s
A1
A2
A3
A4
A5
A6
A7
A8
A9
A10
A11
A12
B1
B2
B3
B4
B5
B6
B7
B8
B9
B10
B11
B12
C1
C2
C3
C4
C5
C6
C7
C8
C9
C10
C11
C12
D1
D2
D3
D4
D5
D6
D7
D8
D9
D10
D11
D12
E1
E2
E3
E4
E5
E6
E7
E8
E9
E10
E11
E12
F1
F2
F3
F4
F5
F6
F7
F8
F9
F10
F11
F12
G1
G2
G3
G4
G5
G6
G7
G8
G9
G10
G11
G12
H1
H2
H3
H4
H5
H6
H7
H8
H9
H10
H11
H12
S A L T R X 1 F O R M U L A T I O N
1. 1.8 M Sodium acetate trihydrate pH 7.0, 0.1 M BIS-TRIS propane pH 7.0
2. 2.8 M Sodium acetate trihydrate pH 7.0, 0.1 M BIS-TRIS propane pH 7.0
3. 1.5 M Ammonium chloride, 0.1 M Sodium acetate trihydrate pH 4.6
4. 1.5 M Ammonium chloride, 0.1 M BIS-TRIS propane pH 7.0
5. 1.5 M Ammonium chloride, 0.1 M Tris pH 8.5
6. 3.5 M Ammonium chloride, 0.1 M Sodium acetate trihydrate pH 4.6
7. 3.5 M Ammonium chloride, 0.1 M BIS-TRIS propane pH 7.0
8. 3.5 M Ammonium chloride, 0.1 M Tris pH 8.5
9. 2.2 M Sodium chloride, 0.1 M Sodium acetate trihydrate pH 4.6
10. 2.2 M Sodium chloride, 0.1 M BIS-TRIS propane pH 7.0
11. 2.2 M Sodium chloride, 0.1 M Tris pH 8.5
12. 3.2 M Sodium chloride, 0.1 M Sodium acetate trihydrate pH 4.6
13. 3.2 M Sodium chloride, 0.1 M BIS-TRIS propane pH 7.0
14. 3.2 M Sodium chloride, 0.1 M Tris pH 8.5
15. 1.0 M Ammonium citrate dibasic, 0.1 M Sodium acetate trihydrate pH 4.6
16. 1.8 M Ammonium citrate dibasic, 0.1 M Sodium acetate trihydrate pH 4.6
17. 1.0 M Ammonium citrate tribasic pH 7.0, 0.1 M BIS-TRIS propane pH 7.0
18. 2.0 M Ammonium citrate tribasic pH 7.0, 0.1 M BIS-TRIS propane pH 7.0
19. 0.7 M Sodium citrate tribasic dihydrate, 0.1 M BIS-TRIS propane pH 7.0
20. 0.7 M Sodium citrate tribasic dihydrate, 0.1 M Tris pH 8.5
21. 1.2 M Sodium citrate tribasic dihydrate, 0.1 M BIS-TRIS propane pH 7.0
22. 1.2 M Sodium citrate tribasic dihydrate, 0.1 M Tris pH 8.5
23. 0.4 M Magnesium formate dihydrate, 0.1 M Sodium acetate trihydrate pH 4.6
24. 0.4 M Magnesium formate dihydrate, 0.1 M BIS-TRIS propane pH 7.0
25. 0.4 M Magnesium formate dihydrate, 0.1 M Tris pH 8.5
26. 0.7 M Magnesium formate dihydrate, 0.1 M BIS-TRIS propane pH 7.0
27. 2.0 M Sodium formate, 0.1 M Sodium acetate trihydrate pH 4.6
28. 2.0 M Sodium formate, 0.1 M BIS-TRIS propane pH 7.0
29. 2.0 M Sodium formate, 0.1 M Tris pH 8.5
30. 3.5 M Sodium formate, 0.1 M Sodium acetate trihydrate pH 4.6
31. 3.5 M Sodium formate, 0.1 M BIS-TRIS propane pH 7.0
32. 3.5 M Sodium formate, 0.1 M Tris pH 8.5
33. 1.2 M DL-Malic acid pH 7.0, 0.1 M BIS-TRIS propane pH 7.0
34. 2.2 M DL-Malic acid pH 7.0, 0.1 M BIS-TRIS propane pH 7.0
35. 1.4 M Sodium malonate pH 7.0, 0.1 M BIS-TRIS propane pH 7.0
36. 2.4 M Sodium malonate pH 7.0, 0.1 M BIS-TRIS propane pH 7.0
37. 2.5 M Ammonium nitrate, 0.1 M Sodium acetate trihydrate pH 4.6
38. 2.5 M Ammonium nitrate, 0.1 M BIS-TRIS propane pH 7.0
39. 2.5 M Ammonium nitrate, 0.1 M Tris pH 8.5
40. 6.0 M Ammonium nitrate, 0.1 M Sodium acetate trihydrate pH 4.6
41. 6.0 M Ammonium nitrate, 0.1 M BIS-TRIS propane pH 7.0
42. 6.0 M Ammonium nitrate, 0.1 M Tris pH 8.5
43. 1.5 M Sodium nitrate, 0.1 M Sodium acetate trihydrate pH 4.6
44. 1.5 M Sodium nitrate, 0.1 M BIS-TRIS propane pH 7.0
45. 1.5 M Sodium nitrate, 0.1 M Tris pH 8.5
46. 4.0 M Sodium nitrate, 0.1 M Sodium acetate trihydrate pH 4.6
47. 4.0 M Sodium nitrate, 0.1 M BIS-TRIS propane pH 7.0
48. 4.0 M Sodium nitrate, 0.1 M Tris pH 8.5
1. 1.0 M Ammonium phosphate monobasic, 0.1 M Sodium acetate trihydrate pH 4.6
2. 1.8 M Ammonium phosphate monobasic, 0.1 M Sodium acetate trihydrate pH 4.6
3. 1.5 M Ammonium phosphate dibasic, 0.1 M Tris pH 8.5
4. 2.4 M Ammonium phosphate dibasic, 0.1 M Tris pH 8.5
5. 1.0 M Sodium phosphate monobasic monohydrate, Potassium phosphate dibasic / pH 5.0
6. 1.0 M Sodium phosphate monobasic monohydrate, Potassium phosphate dibasic / pH 6.9
7. 1.0 M Sodium phosphate monobasic monohydrate, Potassium phosphate dibasic / pH 8.2
8. 1.8 M Sodium phosphate monobasic monohydrate, Potassium phosphate dibasic / pH 5.0
9. 1.8 M Sodium phosphate monobasic monohydrate, Potassium phosphate dibasic / pH 6.9
10. 1.8 M Sodium phosphate monobasic monohydrate, Potassium phosphate dibasic / pH 8.2
11. 0.5 M Succinic acid pH 7.0, 0.1 M BIS-TRIS propane pH 7.0
12. 1.0 M Succinic acid pH 7.0, 0.1 M BIS-TRIS propane pH 7.0
13. 1.5 M Ammonium sulfate, 0.1 M Sodium acetate trihydrate pH 4.6
14. 1.5 M Ammonium sulfate, 0.1 M BIS-TRIS propane pH 7.0
15. 1.5 M Ammonium sulfate, 0.1 M Tris pH 8.5
16. 2.5 M Ammonium sulfate, 0.1 M Sodium acetate trihydrate pH 4.6
17. 2.5 M Ammonium sulfate, 0.1 M BIS-TRIS propane pH 7.0
18. 2.5 M Ammonium sulfate, 0.1 M Tris pH 8.5
19. 0.8 M Lithium sulfate monohydrate, 0.1 M Sodium acetate trihydrate pH 4.6
20. 0.8 M Lithium sulfate monohydrate, 0.1 M BIS-TRIS propane pH 7.0
21. 0.8 M Lithium sulfate monohydrate, 0.1 M Tris pH 8.5e
22. 1.5 M Lithium sulfate monohydrate, 0.1 M Sodium acetate trihydrate pH 4.6
23. 1.5 M Lithium sulfate monohydrate, 0.1 M BIS-TRIS propane pH 7.0
24. 1.5 M Lithium sulfate monohydrate, 0.1 M Tris pH 8.5
25. 1.0 M Magnesium sulfate hydrate, 0.1 M Sodium acetate trihydrate pH 4.6
26. 1.0 M Magnesium sulfate hydrate, 0.1 M BIS-TRIS propane pH 7.0
27. 1.0 M Magnesium sulfate hydrate, 0.1 M Tris pH 8.5
28. 1.8 M Magnesium sulfate hydrate, 0.1 M Sodium acetate trihydrate pH 4.6
29. 1.8 M Magnesium sulfate hydrate, 0.1 M BIS-TRIS propane pH 7.0
30. 1.8 M Magnesium sulfate hydrate, 0.1 M Tris pH 8.5
31. 0.7 M Ammonium tartrate dibasic, 0.1 M Sodium acetate trihydrate pH 4.6
32. 0.7 M Ammonium tartrate dibasic, 0.1 M BIS-TRIS propane pH 7.0
33. 0.7 M Ammonium tartrate dibasic, 0.1 M Tris pH 8.5
34. 1.1 M Ammonium tartrate dibasic, 0.1 M Sodium acetate trihydrate pH 4.6
35. 1.3 M Ammonium tartrate dibasic, 0.1 M BIS-TRIS propane pH 7.0
36. 1.4 M Ammonium tartrate dibasic, 0.1 M Tris pH 8.5
37. 0.6 M Potassium sodium tartrate tetrahydrate 0.1 M BIS-TRIS propane pH 7.0
38. 1.2 M Potassium sodium tartrate tetrahydrate 0.1 M BIS-TRIS propane pH 7.0
39. 0.6 M Potassium sodium tartrate tetrahydrate, 0.1 M Tris pH 8.5
40. 1.2 M Potassium sodium tartrate tetrahydrate, 0.1 M Tris pH 8.5
41. 0.5 M Potassium thiocyanate, 0.1 M Sodium acetate trihydrate pH 4.6
42. 0.5 M Potassium thiocyanate, 0.1 M BIS-TRIS propane pH 7.0
43. 0.5 M Potassium thiocyanate, 0.1 M Tris pH 8.5
44. 4.0 M Ammonium acetate, 0.1 M Sodium acetate trihydrate pH 4.6
45. 0.1 M BIS-TRIS propane pH 7.0, 4.0 M Ammonium Acetate
46. 4.0 M Ammonium acetate, 0.1 M Tris pH 8.5
47. 35% v/v Tacsimate

, 0.1 M BIS-TRIS propane pH 7.0

48. 60% v/v Tacsimate

, 0.1 M BIS-TRIS propane pH 7.0

S A L T R X 2 F O R M U L A T I O N
Ammonium citrate dibasic
Ammonium citrate tribasic
Magnesium formate dihydrate
Sodium acetate
Sodium citrate tribasic dihydrate
Sodium formate
Sodium malonate
DL-Malic acid
Range from 4.6 to 8.5
p H
o r g a n i c s a l t
Ammonium chloride
Sodium chloride
Ammonium nitrate
Sodium nitrate
s a l t
BIS-TRIS propane
Sodium acetate trihydrate
Tris
Ammonium acetate
Ammonium tartrate dibasic
K/Na Tartrate
Succinic acid
Tacsimate
TM
Range from 4.6 to 8.5
buffer
p H
o r g a n i c s a l t
Ammonium phosphate mono.
Ammonium phosphate dibasic
Ammonium sulfate
Lithium sulfate monohydrate
Magnesium sulfate hydrate
Na/K Phosphate
Potassium thiocyanate
s a l t
BIS-TRIS propane
Sodium acetate trihydrate
Tris
buffer
s a l t r x 1
f a c t o r s
s a l t r x 2
f a c t o r s
Crystals above are those of the membrane protein
succinate:quinone oxireductase. Preliminary crystalliza-
tion conditions determined using MembFac.
Courtesy of Michael Stowell.
MRC-LMB
s u c c e s s s t o r y
n
Primary sparse matrix crystallization screen
for membrane proteins and samples with
limited solubility
n
Membrane protein sparse matrix screens
1
n
Sparse matrix formula efficiently samples
salts, polymers, organics and pH
n
pH range 4.6 - 8.5
n
Formulated for use with detergents
n
Crystal Screen Lite features a lower ionic
strength than the original Crystal Screen
n
Tube or Deep Well block format
a p p l i c a t i o n
f e a t u r e s
MembFac is a highly effective sparse matrix
screen specifically designed as a preliminary
screen for the crystallization ofmembrane pro-
teins. MembFac is based upon the highly effec-
tive biased sparse matrix methodology. The
MembFac screen matrix is optimized for the
crystallization of membrane proteins.
MembFac contains 48 unique reagent formula-
tions. With this set of 48 conditions, numer-
ous chemicals and chemical combinations are
utilized, allowing one to evaluate a large variety of potential crystallization conditions. MembFac covers a
broad pH range between 4 and 9.
The Rationale
The basic rationale of MembFac is to perform a sparse matrix screen while changing the detergent dimen-
sion because detergents play an important factor in the crystallization of membrane proteins. A MembFac
screen is to be completed for each detergent. Detergent screens are available separately.
The Method
The membrane protein of interest is first isolated in the detergent which gives the highest stability and
activity. The protein concentration should be approximately 10 to 20 mg/ml and the detergent concentra-
tion should be only slightly above the CMC. The sample is then set against MembFac, using all 48 solutions
plus the screening detergent(s) of choice. The screening detergent concentration should be approximately
one to three times the CMC. Hence, for each detergent screened, one would utilize approximately 1 to 2
mg of sample.
The MembFac protocol was used to crystallize membrane proteins including SQR, fumerate: quinone
oxireductase, LHII, and LHCII.
Crystal Screen Lite is a sparse matrix of trial crystallization reagent conditions based upon the Crystal
Screen.
3
The primary screen variables are salt, pH, and precipitant (salts, polymers, volatile organics, and
non-volatile organics).
2
Crystal Screen Lite differs from the original Crystal Screen kit in that the primary
precipitant reagents are one-half the concentration of that used in the original Crystal Screen formulation.
The secondary salts, ions, and buffers remain at the original Crystal Screen concentration. Reducing the
primary concentration of the primary precipitant results in a screen which is more gentle on the sample
and typically produces much less precipitate conditions than the original Crystal Screen.
MembFac contains 48 unique reagents, 10 ml each.
MembFac HT contains 1 ml each of all 48 reagents from MembFac and reagents 1-48 from Crystal Screen
Lite in a single Deep Well block format.
Ready-to-use reagents are sterile filtered and formulated with ultra-pure Type 1 water, using the highest
purity salts, polymers, organics and buffers.
References
1. UCLA Crystallization Workshop, June 21, 1993.
2. Crystallization of nucleic acids and proteins, Edited by A. Ducruix and R. Giege, The Practical Approach Series, Oxford Univ. Press, 1992.
3. McPherson, A., Current approaches to macromolecular crystallization., Eur. J. Biochem. (1990) 189, 1-23.
4. Jancarik, J. and Kim, S.H., Sparse Matrix Sampling: a screening method for crystallization of proteins. (1991) J. Appl. Cryst., 24,409-411.
5. Protein and Nucleic Acid Crystallization. Methods, A Companion to Methods in Enzymology, Academic Press, Volume 1, Number 1, August 1990.
Order Information
Each MembFac kit contains 48 unique reagents. Crystal Screen Lite kit contains 50 unique
reagents. To order individual reagents, use Custom Shop

catalog number listed below. Refer to

page 36 for further details.
Cat. No. Name Description Price
HR2-114 MembFac 10 ml, tube format $285.00
HR2-128 Crystal Screen Lite 10 ml, tube format $285.00
HR2-137 MembFac HT 1 ml, Deep Well block format $185.00
HR2-920-** MembFac Custom Shop 185 ml $138.00
HR2-916-** Crystal Screen Lite Custom Shop 185 ml $138.00
** = reagent number 1-48 (for MembFac)
** = reagent number 1-50 (for Crystal Screen Lite)
d e s c r i p t i o n
Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com 18
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M E M B F A C

C r y s T a l s C r e e n l I T e

M e M b F a C H T

19
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Range from 4.6 to 8.5
2-Propanol
(+/-)-2-Methyl-2,4-pentanediol
Polyethylene glycol 400
Polyethylene glycol 4,000
Polyethylene glycol 6,000
Magnesium chloride
Sodium chloride
Ammonium sulfate
Lithium sulfate
Magnesium sulfate
Ammonium phosphate
Potassium phosphate
Sodium phosphate
Sodium citrate
K/Na tartrate
o r g a n i c
p
H
s a l t
p o l y m e r
n o n - v o l a t i l e
o r g a n i c
A1
A2
A3
A4
A5
A6
A7
A8
A9
A10
A11
A12
B1
B2
B3
B4
B5
B6
B7
B8
B9
B10
B11
B12
C1
C2
C3
C4
C5
C6
C7
C8
C9
C10
C11
C12
D1
D2
D3
D4
D5
D6
D7
D8
D9
D10
D11
D12
E1
E2
E3
E4
E5
E6
E7
E8
E9
E10
E11
E12
F1
F2
F3
F4
F5
F6
F7
F8
F9
F10
F11
F12
G1
G2
G3
G4
G5
G6
G7
G8
G9
G10
G11
G12
H1
H2
H3
H4
H5
H6
H7
H8
H9
H10
H11
H12
1. 0.1 M Sodium chloride, 0.1 M Sodium acetate trihydrate pH 4.6, 12% v/v (+
/
-)-2-Methyl-2,4-pentanediol
2. 0.1 M Zinc acetate dihydrate, 0.1 M Sodium acetate trihydrate pH 4.6, 12% w/v Polyethylene glycol 4,000
3. 0.2 M Ammonium sulfate, 0.1 M Sodium acetate trihydrate pH 4.6, 10% w/v Polyethylene glycol 4,000
4. 0.1 M Sodium chloride, 0.1 M Sodium acetate trihydrate pH 4.6, 12% v/v 2-Propanol
5. 0.1 M Sodium acetate trihydrate pH 4.6, 12% w/v Polyethylene glycol 4,000
6. 0.1 M Sodium acetate trihydrate pH 4.6, 1.0 M Ammonium sulfate
7. 0.1 M Sodium acetate trihydrate pH 4.6, 1.0 M Magnesium sulfate heptahydrate
8. 0.1 M Magnesium chloride hexahydrate, 0.1 M Sodium acetate trihydrate pH 4.6, 18% v/v Polyethylene glycol 400
9. 0.1 M Lithium sulfate monohydrate, 0.1 M Sodium acetate trihydrate pH 4.6, 1.0 M Ammonium phosphate monobasic
10. 0.1 M Sodium chloride, 0.1 M Sodium acetate trihydrate pH 4.6, 12% w/v Polyethylene glycol 6,000
11. 0.1 M Magnesium chloride hexahydrate, 0.1 M Sodium acetate trihydrate pH 4.6, 12% w/v Polyethylene glycol 6,000
12. 0.1 M Sodium chloride, 0.1 M Sodium citrate tribasic dihydrate pH 5.6, 18% v/v Polyethylene glycol 400
13. 0.1 M Lithium sulfate monohydrate, 0.1 M Sodium citrate tribasic dihydrate pH 5.6, 12 % w/v Polyethylene glycol 4,000
14. 0.1 M Sodium citrate tribasic dihydrate, 0.1 M Sodium citrate tribasic dihydrate pH 5.6, 10% v/v 2-Propanol
15. 0.1 M Sodium chloride, 0.1 M Sodium citrate tribasic dihydrate pH 5.6, 12% v/v (+
/
-)-2-Methyl-2,4-pentanediol
16. 0.1 M Sodium citrate tribasic dihydrate pH 5.6, 1.0 M Magnesium sulfate heptahydrate
17. 0.1 M Sodium chloride, 0.1 M Sodium citrate tribasic dihydrate pH 5.6, 12% w/v Polyethylene glycol 4,000
18. 0.1 M Lithium sulfate monohydrate, 0.1 M Sodium citrate tribasic dihydrate pH 5.6, 12% w/v Polyethylene glycol 6,000
19. 0.1 M Magnesium chloride hexahydrate, 0.1 M Sodium citrate tribasic dihydrate pH 5.6, 4% v/v (+
/
-)-2-Methyl-2,4-pentanediol
20. 0.1 M Sodium citrate trihydrate dihydrate pH 5.6, 0.1 M Sodium chloride
21. 0.1 M Lithium sulfate monohydrate, 0.1 M Sodium citrate tribasic dihydrate pH 5.6, 4% v/v Polyethylene glycol 400
22. 0.1 M ADA pH 6.5, 1.0 M Ammonium sulfate
23. 0.1 M Lithium sulfate monohydrate, 0.1 M ADA pH 6.5, 12% w/v Polyethylene glycol 4,000, 2% v/v 2-Propanol
24. 0.1 M ADA pH 6.5, 1.0 M Ammonium phosphate dibasic
25. 0.1 M Magnesium chloride hexahydrate, 0.1 M ADA pH 6.5, 12% w/v Polyethylene glycol 6,000
26. 0.1 M ADA pH 6.5, 12% v/v (+
/
-)-2-Methyl-2,4-pentanediol
27. 0.1 M Lithium sulfate monohydrate, 0.1 M ADA pH 6.5, 1.0 M Magnesium sulfate hydrate
28. 0.3 M Lithium sulfate monohydrate, 0.1 M ADA pH 6.5, 4% v/v Polyethylene glycol 400
29. 0.1 M Ammonium sulfate, 0.1 M HEPES sodium pH 7.5, 0.5 M Sodium phosphate dibasic dihydrate,
0.5 M Potassium phosphate dibasic
30. 0.1 M Sodium chloride, 0.1 M HEPES sodium pH 7.5, 10% w/v Polyethylene glycol 4,000
31. 0.1 M Magnesium chloride hexahydrate, 0.1 M HEPES sodium pH 7.5, 18% v/v Polyethylene glycol 400
32. 0.1 M HEPES sodium pH 7.5, 1.0 M Potassium sodium tartrate tetrahydrate
33. 0.1 M Ammonium sulfate, 0.1 M HEPES sodium pH 7.5, 18% v/v Polyethylene glycol 400
34. 0.1 M Ammonium sulfate, 0.1 M HEPES sodium pH 7.5, 10% w/v Polyethylene glycol 4,000
35. 0.1 M Sodium citrate tribasic dihydrate, 0.1 M HEPES sodium pH 7.5, 12% v/v (+
/
-)-2-Methyl-2,4-pentanediol
36. 0.1 M HEPES sodium pH 7.5, 1.0 M Sodium citrate tribasic dihydrate
37. 0.6 M Magnesium sulfate hydrate, 0.1 M HEPES sodium pH 7.5, 4% v/v Polyethylene glycol 400
38. 0.6 M Magnesium sulfate hydrate, 0.1 M HEPES sodium pH 7.5, 4% v/v (+
/
-)-2-Methyl-2,4-pentanediol
39. 0.1 M Lithium sulfate monohydrate, 0.1 M HEPES sodium pH 7.5, 0.1 M Potassium sodium tartrate tetrahydrate
40. 0.1 M Lithium sulfate monohydrate, 0.1 M TRIS hydrochloride pH 8.5, 12% v/v (+
/
-)-2-Methyl-2,4-pentanediol
41. 0.1 M Ammonium phosphate dibasic, 0.1 M TRIS hydrochloride pH 8.5, 0.5 M Sodium phosphate dibasic dihydrate,
0.5 M Potassium phosphate dibasic
42. 0.1 M TRIS hydrochloride pH 8.5, 0.1 M Sodium acetate trihydrate
43. 0.1 M TRIS hydrochloride pH 8.5, 0.1 M Sodium chloride
44. 0.1 M Ammonium phosphate dibasic, 0.1 M TRIS hydrochloride pH 8.5, 12% w/v Polyethylene glycol 6,000
45. 0.1 M Potassium sodium tartrate tetrahydrate, 0.1 M TRIS hydrochloride pH 8.5, 0.4 M Magnesium sulfate hydrate
46. 0.1 M TRIS hydrochloride pH 8.5, 0.2 M Lithium sulfate monohydrate
47. 0.1 M TRIS hydrochloride pH 8.5, 0.5 M Ammonium sulfate
48. 0.1 M Sodium citrate tribasic dihydrate, 0.1 M TRIS hydrochloride pH 8.5, 5% v/v Polyethylene glycol 400
1. 0.02 M Calcium chloride dihydrate, 0.1 M Sodium acetate trihydrate pH 4.6, 15% v/v (+
/
-)-2-Methyl-2,4-pentanediol
2. 0.2 M Potassium sodium tartrate tetrahydrate
3. 0.2 M Ammonium phosphate monobasic
4. 0.1 M TRIS hydrochloride pH 8.5, 1.0 M Ammonium sulfate
5. 0.2 M Sodium citrate tribasic dihydrate, 0.1 M HEPES sodium pH 7.5, 15% v/v (+
/
-)-2-Methyl-2,4-pentanediol
6. 0.2 M Magnesium chloride hexahydrate, 0.1 M TRIS hydrochloride pH 8.5, 15% w/v Polyethylene glycol 4,000
7. 0.1 M Sodium cacodylate trihydrate pH 6.5, 0.7 M Sodium acetate trihydrate
8. 0.2 M Sodium citrate tribasic dihydrate, 0.1 M Sodium cacodylate trihydrate pH 6.5, 15% v/v 2-Propanol
9. 0.2 M Ammonium acetate, 0.1 M Sodium citrate tribasic dihydrate pH 5.6, 15% w/v Polyethylene glycol 4,000
10. 0.2 M Ammonium acetate, 0.1 M Sodium acetate trihydrate pH 4.6, 15% w/v Polyethylene glycol 4,000
11. 0.1 M Sodium citrate tribasic dihydrate pH 5.6, 0.5 M Ammonium phosphate monobasic
12. 0.2 M Magnesium chloride hexahydrate, 0.1 M HEPES sodium pH 7.5, 15% v/v 2-Propanol
13. 0.2 M Sodium citrate tribasic dihydrate, 0.1 M TRIS hydrochloride pH 8.5, 15% v/v Polyethylene glycol 400
14. 0.2 M Calcium chloride dihydrate, 0.1 M HEPES sodium pH 7.5, 14% v/v Polyethylene glycol 400
15. 0.2 M Ammonium sulfate, 0.1 M Sodium cacodylate trihydrate pH 6.5, 15% w/v Polyethylene glycol 8,000
16. 0.1 M HEPES sodium pH 7.5, 0.75 M Lithium sulfate monohydrate
17. 0.2 M Lithium sulfate monohydrate, 0.1 M TRIS hydrochloride pH 8.5, 15% w/v Polyethylene glycol 4,000
18. 0.2 M Magnesium acetate tetrahydrate, 0.1 M Sodium cacodylate trihydrate pH 6.5, 10% w/v Polyethylene glycol 8,000
19. 0.2 M Ammonium acetate, 0.1 M TRIS hydrochloride pH 8.5, 15% v/v 2-Propanol
20. 0.2 M Ammonium sulfate, 0.1 M Sodium acetate trihydrate pH 4.6, 12.5% w/v Polyethylene glycol 4,000
21. 0.2 M Magnesium acetate tetrahydrate, 0.1 M Sodium cacodylate trihydrate pH 6.5, 15% v/v (+
/
-)-2-Methyl-2,4-pentanediol
22. 0.2 M Sodium acetate trihydrate, 0.1 M TRIS hydrochloride pH 8.5, 15% w/v Polyethylene glycol 4,000
23. 0.2 M Magnesium chloride hexahydrate, 0.1 M HEPES sodium pH 7.5, 15% v/v Polyethylene glycol 400
24. 0.2 M Calcium chloride dihydrate, 0.1 M Sodium acetate trihydrate pH 4.6, 10% v/v 2-Propanol
25. 0.1 M Imidazole pH 6.5, 0.5 M Sodium acetate trihydrate
26. 0.2 M Ammonium acetate, 0.1 M Sodium citrate tribasic dihydrate pH 5.6, 15% v/v (+
/
-)-2-Methyl-2,4-pentanediol
27. 0.2 M Sodium citrate tribasic dihydrate, 0.1 M HEPES sodium pH 7.5, 10% v/v 2-Propanol
28. 0.2 M Sodium acetate trihydrate, 0.1 M Sodium cacodylate trihydrate pH 6.5, 15% w/v Polyethylene glycol 8,000
29. 0.1 M HEPES sodium pH 7.5, 0.4 M Potassium sodium tartrate tetrahydrate
30. 0.2 M Ammonium sulfate, 15% w/v Polyethylene glycol 8,000
31. 0.2 M Ammonium sulfate, 15% w/v Polyethylene glycol 4,000
32. 1.0 M Ammonium sulfate
33. 2.0 M Sodium formate
34. 0.1 M Sodium acetate trihydrate pH 4.6, 1.0 M Sodium formate
35. 0.1 M HEPES sodium pH 7.5, 0.4 M Sodium phosphate monobasic monohydrate, 0.4 M Potassium phosphate monobasic
36. 0.1 M TRIS hydrochloride pH 8.5, 4% w/v Polyethylene glycol 8,000
37. 0.1 M Sodium acetate trihydrate pH 4.6, 4% w/v Polyethylene glycol 4,000
38. 0.1 M HEPES sodium pH 7.5, 0.7 M Sodium citrate tribasic dihydrate
39. 0.1 M HEPES sodium pH 7.5, 2% v/v Polyethylene glycol 400, 1.0 M Ammonium sulfate
40. 0.1 M Sodium citrate tribasic dihydrate pH 5.6, 10% v/v 2-Propanol, 10% w/v Polyethylene glycol 4,000
41. 0.1 M HEPES sodium pH 7.5, 5% v/v 2-Propanol, 10% w/v Polyethylene glycol 4,000
42. 0.05 M Potassium phosphate monobasic, 10% w/v Polyethylene glycol 8,000
43. 15% w/v Polyethylene glycol 1,500
44. 0.1 M Magnesium formate dihydrate
45. 0.2 M Zinc acetate dihydrate, 0.1 M Sodium cacodylate trihydrate pH 6.5, 9% w/v Polyethylene glycol 8,000
46. 0.2 M Calcium acetate hydrate, 0.1 M Sodium cacodylate trihydrate pH 6.5, 9% w/v Polyethylene glycol 8,000
47. 0.1 M Sodium acetate trihydrate pH 4.6, 1.0 M Ammonium sulfate
48. 0.1 M TRIS hydrochloride pH 8.5, 1.0 M Ammonium phosphate monobasic
49. 0.5 M Lithium sulfate monohydrate, 2% w/v Polyethylene glycol 8,000
50.

0.5 M Lithium sulfate monohydrate, 7.5% w/v Polyethylene glycol 8,000
C R Y S T A L S C R E E N L I T E F O R M U L A T I O N
M E M B F A C F O R M U L A T I O N
Range from 4.0 to 9.0
2-Propanol
(+/-)-2-Methyl-2,4-pentanediol
Polyethylene glycol 400
Polyethylene glycol 1,500
Polyethylene glycol 4,000
Polyethylene glycol 8,000
Ammonium acetate
Calcium acetate
Magnesium acetate
Sodium acetate
Zinc acetate
Calcium chloride
Magnesium chloride
Sodium citrate
Magnesium formate
Sodium formate
Ammonium phosphate
Potassium phosphate
Sodium phosphate
Ammonium sulfate
Lithium sulfate
K/Na tartrate
o r g a n i c
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s a l t
p o l y m e r
HEPES sodium
Imidazole
Sodium acetate
Sodium cacodylate
Sodium citrate
TRIS hydrochloride
b u f f e r
m e m b f a c
f a c t o r s
crystal screen lite
f a c t o r s
n
Crystallization screen for nucleic acids &
protein/nucleic acid complexes
n
Nucleic acid sparse matrix screen
1
n
Sparse matrix formula efficiently samples
salts, polyols, organics, & pH
n
pH range 5.6 - 8.5
n
Screen Magnesium, Potassium,
& Ammonium anions
a p p l i c a t i o n
f e a t u r e s
Natrix is based upon a published protocol
which is highly effective for the crystallization
of nucleic acids and protein-nucleic acid com-
plexes. A variety of hammerhead ribozymes
and other ribozymes, RNAs, DNAs, RNA-drug
complexes, and RNA-protein complexes have
been crystallized using the Natrix protocol.
By using sparse matrix sampling technology,
Natrix allows one to quickly test wide ranges
of pH, salts, and precipitants using a very small sample (50 to 100 l) of nucleic acid.
Natrix is unique in that rather than relying solely on the traditional nucleic acid precipitant MPD, Natrix
also utilizes Polyethylene glycols (PEGs) in a variety of molecular weights (200, 400, 4,000, 8,000) as well as
2-Propanol, Polyethylene glycol monomethyl ether (PEG MME), and 1,6-Hexanediol. Many of the polymeric
and low molecular weight organic precipitants are combined with various monovalent salts as precipitating
agents. This combination of salts and low molecular weight organics and polyalcohols, as well as the utiliza-
tion of varying chain length PEGs, has proven to be a successful combination for producing nucleic acid
and protein-nucleic acid complex crystals.
RNA Crystallization
Typical crystallization trials using Natrix will use the hanging or sitting drop methodology. Sample prepara-
tion is straightforward. The highly purified sample is heated to 75C and annealed in the presence of the
appropriate polyamine and buffer. After cooling to room temperature, the sample is mixed with an equal
amount of Natrix solution and set against a reservoir containing the same Natrix solution for vapor diffusion
equilibration. Typically, crystals are observed within the first week of setting up the trial.
Each Natrix contains 48 unique reagents of varying salt and precipitant composition over six different
pH levels. Each preformulated, sterile filtered solution provides sufficient reagent for more than a dozen
screens. The method is suitable for hanging drop, sitting drop, sandwich drop, and microbatch crystalliza-
tion methodologies. Crystallization plates are sold separately. Natrix does not include polyamines. It is rec-
ommended that a polyamine such as Spermine be added to the sample prior to screening and that various
polyamines be screened AFTER preliminary crystallization conditions have been determined.
Each Natrix kit contains 48 unique reagents, 10 ml each. Ready-to-use reagents are sterile filtered and
formulated with ultra-pure Type 1 water, using the highest purity salts, polymers, organics, and buffers.
References
1. Scott, W. G., Finch, J.T., Grenfell, R., Fogg, J., Smith, T., Gait, M.J., Klug, A., Journal of Molecular Biology (1995) 250: 327-332.
2. Purification, crystallization and preliminary X-ray analysis of the BseCI DNA methyltransferase from Bacillus stearothermophilus in complex with it's cognate
DNA. E. G. Kapetaniou, D. Kotsifaki, M. Providaki, M. Rina, V. Bouriotis and M. Kokkinidis. Acta Cryst. (2007). F63, 12-14.
3. Crystallization and preliminary X-ray diffraction analysis of an Escherichia coli tRNAGly acceptor-stem microhelix. C. Frster, M. Perbandt, A. B. E. Brauer,
S. Brode, J. P. Frste, C. Betzel and V. A. Erdmann. Acta Cryst. (2007). F63, 46-48.
4. Combinatorial crystallization of an RNA-protein complex. Danielle Bodrero Hoggan, Jeffrey A. Chao, G. S. Prasad, C. David Stouta and James R. Williamson.
Acta Cryst (2003). D59, 466-473
5. Cocrystallizing natural RNA with its unnatural mirror image: biochemical and preliminary X-ray diffraction analysis of a 5S rRNA A-helix racemate.
Volker A. Erdmanna et al. Acta Cryst. (2007). F63, 839843.
Order Information
Each Natrix kit contains 48 unique reagents. To order individual reagents, use Custom Shop

catalog number listed below. Refer to page 36 for further details.

Cat. No. Name Description Price
HR2-116 Natrix 10 ml, tube format $285.00
HR2-918-** Natrix Custom Shop 185 ml $138.00
** = reagent number 1-48
d e s c r i p t i o n
Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com 20
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Crystal of MtaN, the N-terminal DNA binding domain of
the Multidrug Transporter Activation protein from B. subti-
lis. Initial crystals grown from Natrix reagent number 29.
Courtesy of Michael Godsey.
Oregon Health Sciences University, Dept. of Biochemistry
and Molecular Biology
s u c c e s s s t o r y
N A T R I X

21
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1. 0.01 M Magnesium chloride hexahydrate, 0.05 M MES monohydrate pH 5.6, 1.8 M Lithium sulfate monohydrate
2. 0.01 M Magnesium acetate tetrahydrate, 0.05 M MES monohydrate pH 5.6, 2.5 M Ammonium sulfate
3. 0.1 M Magnesium acetate tetrahydrate, 0.05 M MES monohydrate pH 5.6,
20% v/v (+
/
-)-2-Methyl-2,4-pentanediol
4. 0.2 M Potassium chloride, 0.01 M Magnesium sulfate heptahydrate, 0.05 M MES monohydrate pH 5.6,
10% v/v Polyethylene glycol 400
5. 0.2 M Potassium chloride, 0.01 M Magnesium chloride hexahydrate, 0.05 M MES monohydrate pH 5.6,
5% w/v Polyethylene glycol 8,000
6. 0.1 M Ammonium sulfate, 0.01 M Magnesium chloride hexahydrate, 0.05 M MES monohydrate pH 5.6,
20% w/v Polyethylene glycol 8,000
7. 0.02 M Magnesium chloride hexahydrate, 0.05 M MES monohydrate pH 6.0, 15% v/v 2-Propanol
8. 0.1 M Ammonium acetate, 0.005 M Magnesium sulfate heptahydrate, 0.05 M MES monohydrate pH 6.0,
0.6 M Sodium chloride
9. 0.1 M Potassium chloride, 0.01 M Magnesium chloride hexahydrate, 0.05 M MES monohydrate pH 6.0,
10% v/v Polyethylene glycol 400
10. 0.005 M Magnesium sulfate heptahydrate, 0.05 M MES monohydrate pH 6.0, 5% w/v Polyethylene glycol 4,000
11. 0.01 M Magnesium chloride hexahydrate, 0.05 M Sodium cacodylate trihydrate pH 6.0,
1.0 M Lithium sulfate monohydrate
12. 0.01 M Magnesium sulfate heptahydrate, 0.05 M Sodium cacodylate trihydrate pH 6.0,
1.8 M Lithium sulfate monohydrate
13. 0.015 M Magnesium acetate tetrahydrate, 0.05 M Sodium cacodylate trihydrate pH 6.0, 1.7 M Ammonium sulfate
14. 0.1 M Potassium chloride, 0.025 M Magnesium chloride hexahydrate,
0.05 M Sodium cacodylate trihydrate pH 6.0, 15% v/v 2-Propanol
15. 0.04 M Magnesium chloride hexahydrate, 0.05 M Sodium cacodylate trihydrate pH 6.0,
5% v/v (+
/
-)-2-Methyl-2,4-pentanediol
16. 0.04 M Magnesium acetate tetrahydrate, 0.05 M Sodium cacodylate trihydrate pH 6.0,
30% v/v (+
/
-)-2-Methyl-2,4-pentanediol
17. 0.2 M Potassium chloride, 0.01 M Calcium chloride dihydrate, 0.05 M Sodium cacodylate trihydrate pH 6.0,
10% w/v Polyethylene glycol 4,000
18. 0.01 M Magnesium acetate tetrahydrate, 0.05 M Sodium cacodylate trihydrate pH 6.5,
1.3 M Lithium sulfate monohydrate
19. 0.01 M Magnesium sulfate heptahydrate, 0.05 M Sodium cacodylate trihydrate pH 6.5, 2.0 M Ammonium sulfate
20. 0.1 M Ammonium acetate, 0.015 M Magnesium acetate tetrahydrate, 0.05 M Sodium cacodylate trihydrate pH 6.5,
10% v/v 2-Propanol
21. 0.2 M Potassium chloride, 0.005 M Magnesium chloride hexahydrate, 0.05 M Sodium cacodylate trihydrate pH 6.5,
0.9 M 1,6-Hexanediol
22. 0.08 M Magnesium acetate tetrahydrate, 0.05 M Sodium cacodylate trihydrate pH 6.5,
15% v/v Polyethylene glycol 400
23. 0.2 M Potassium chloride, 0.01 M Magnesium chloride hexahydrate, 0.05 M Sodium cacodylate trihydrate pH 6.5,
10% w/v Polyethylene glycol 4,000
24. 0.2 M Ammonium acetate, 0.01 M Calcium chloride dihydrate, 0.05 M Sodium cacodylate trihydrate pH 6.5,
10% w/v Polyethylene glycol 4,000
25. 0.08 M Magnesium acetate tetrahydrate, 0.05 M Sodium cacodylate trihydrate pH 6.5,
30% w/v Polyethylene glycol 4,000
26. 0.2 M Potassium chloride, 0.1 M Magnesium acetate tetrahydrate, 0.05 M Sodium cacodylate trihydrate pH 6.5,
10 % w/v Polyethylene glycol 8,000
27. 0.2 M Ammonium acetate, 0.01 M Magnesium acetate tetrahydrate, 0.05 M Sodium cacodylate trihydrate pH 6.5,
30% w/v Polyethylene glycol 8,000
28. 0.05 M Magnesium sulfate hydrate, 0.05 M HEPES sodium pH 7.0, 1.6 M Lithium sulfate monohydrate
29. 0.01 M Magnesium chloride hexahydrate, 0.05 M HEPES sodium pH 7.0, 4.0 M Lithium chloride
30. 0.01 M Magnesium chloride hexahydrate, 0.05 M HEPES sodium pH 7.0, 1.6 M Ammonium sulfate
31. 0.005 M Magnesium chloride hexahydrate, 0.05 M HEPES sodium pH 7.0,
25% v/v Polyethylene glycol monomethyl ether 550
32. 0.2 M Potassium chloride, 0.01 M Magnesium chloride hexahydrate, 0.05 M HEPES sodium pH 7.0,
1.7 M 1,6-Hexanediol
33. 0.2 M Ammonium chloride, 0.01 M Magnesium chloride hexahydrate, 0.05 M HEPES sodium pH 7.0,
2.5 M 1,6-Hexanediol
34. 0.1 M Potassium chloride, 0.005 M Magnesium sulfate hydrate, 0.05 M HEPES sodium pH 7.0,
15% v/v (+
/
-)-2-Methyl-2,4-pentanediol
35. 0.1 M Potassium chloride, 0.01 M Magnesium chloride hexahydrate, 0.05 M HEPES sodium pH 7.0,
5% v/v Polyethylene glycol 400
36. 0.1 M Potassium chloride, 0.01 M Calcium chloride dihydrate, 0.05 M HEPES sodium pH 7.0,
10% v/v Polyethylene glycol 400
37. 0.2 M Potassium chloride, 0.025 M Magnesium sulfate hydrate, 0.05 M HEPES sodium pH 7.0,
20% v/v Polyethylene glycol 200
38. 0.2 M Ammonium acetate, 0.15 M Magnesium acetate tetrahydrate, 0.05 M HEPES sodium pH 7.0,
5% w/v Polyethylene glycol 4,000
39. 0.1 M Ammonium acetate, 0.02 M Magnesium chloride hexahydrate, 0.05 M HEPES sodium pH 7.0,
5% w/v Polyethylene glycol 8,000
40. 0.01 M Magnesium chloride hexahydrate, 0.05 M TRIS hydrochloride pH 7.5, 1.6 M Ammonium sulfate
41. 0.1 M Potassium chloride, 0.015 M Magnesium chloride hexahydrate, 0.05 M TRIS hydrochloride pH 7.5
10% v/v Polyethylene glycol monomethyl ether 550
42. 0.01 M Magnesium chloride hexahydrate, 0.05 M TRIS hydrochloride pH 7.5, 5% v/v 2-Propanol
43. 0.05 M Ammonium acetate, 0.01 M Magnesium chloride hexahydrate, 0.05 M TRIS hydrochloride pH 7.5,
10% v/v (+
/
-)-2-Methyl-2,4-pentanediol
44. 0.2 M Potassium chloride, 0.05 M Magnesium chloride hexahydrate, 0.05 M TRIS hydrochloride pH 7.5,
10% w/v Polyethylene glycol 4,000
45. 0.025 M Magnesium sulfate hydrate, 0.05 M TRIS hydrochloride pH 8.5, 1.8 M Ammonium sulfate
46. 0.005 M Magnesium sulfate hydrate, 0.05 M TRIS hydrochloride pH 8.5, 2.9 M 1,6-Hexanediol
47. 0.1 M Potassium chloride, 0.01 M Magnesium chloride hexahydrate, 0.05 M TRIS hydrochloride pH 8.5,
30% v/v Polyethylene glycol 400
48. 0.2 M Ammonium chloride, 0.01 M Calcium chloride dihydrate, 0.05 M TRIS hydrochloride pH 8.5,
30% w/v Polyethylene glycol 4,000
N A T R I X F O R M U L A T I O N
Range from 5.6 to 8.5
p
H
o r g a n i c
p o l y m e r
n o n - v o l a t i l e
o r g a n i c
s a l t
(+/-)-2-Methyl-2,4-pentanediol
1,6-Hexanediol
2-Propanol
Ammonium acetate
Ammonium chloride
Ammonium sulfate
Calcium chloride
Lithium chloride
Lithium sulfate
Magnesium chloride
Magnesium acetate
Magnesium sulfate
Potassium chloride
Polyethylene glycol 200
Polyethylene glycol 400
Polyethylene glycol 4,000
Polyethylene glycol 8,000
Polyethylene glycol MME 550
n a t r i x
f a c t o r s
n
Crystallization screen for proteins, peptides,
& nucleic acids
n
Cryo ready reagents for cryocrystallography
1
n
Sparse matrix formula efficiently samples
salts, polymers, organics, & pH
n
Screen for cryo and crystallization conditions
using only one screen
a p p l i c a t i o n
f e a t u r e s
m e t h o d o l o g i e s
The proportion of macromolecular x-ray
structures solved using cryocrystallographic
methods is increasing at an exponential rate.
The advantages of macromolecular cryocrys-
tallography are numerous. The most widely
used method of flash cooling macromolecu-
lar crystals is to support the crystal in a film of
cryoprotected mother liquor on or in a small
fiber loop which is subsequently cooled in
liquid nitrogen at around 100K. The liquid about the crystal must freeze as an amorphous glass to avoid
crystal damage and diffraction from ordered ice. Cryoprotection is achieved by mixing the mother liquor
and crystal with a suitable cryoprotectant reagent. The most frequently utilized cryoprotectant is glycerol.
Determining a Cryoprotectant
Determining initial and optimal cryoprotectant concentration is often a process of trial and error. One must
find suitable cryoprotectant concentrations which stabilize the crystal while at the same time combine with
the crystallization reagent to form an amorphous glass.
Crystal Screen Cryo removes the guesswork from determining the preliminary glycerol concentration
required to mix with the crystallization reagent to form an amorphous glass. Crystal Screen Cryo reagents
are preformulated with the appropriate amount of glycerol required to form an amorphous glass with
each unique reagent composition. Crystals obtained in Crystal Screen Cryo will already have a suitable
concentration of glycerol in the reagent. This avoids exhaustive empirical searches for preliminary cryopro-
tectant conditions. From the preliminary cryoprotectant conditions, cryoprotectant concentration can be
optimized to minimize mosaic spread and maximize diffraction resolution.
Crystal Screen Cryo is a complete sparse matrix reagent kit designed to provide a rapid screening method
for the crystallization of biological macromolecules in the presence of glycerol, the most frequently utilized
cryoprotectant. Crystal Screen Cryo utilizes the original Crystal Screen protocol but is optimized to
include the appropriate concentration of glycerol required to form an amorphous glass at 100K. The pri-
mary screen variables are salt, pH, precipitant (salts, polymers, volatile organics, and non-volatile organics),
and cryoprotectant concentration. The screen is a straightforward, effective, and practical kit for determin-
ing preliminary crystallization conditions and provides a good starting point for finding suitable cryopro-
tectant conditions for macromolecular crystals grown in a wide range of reagents. Crystal Screen Cryo is
also effective in determining the solubility of a macromolecule in a wide range of precipitants and pH.
Each Crystal Screen Cryo kit contains 50 unique reagents, 10 ml each. Ready-to-use reagents are sterile
filtered and formulated with ultra-pure Type 1 water, using the highest purity salts, polymers, organics,
and buffers.
References
1. Garman, E.F. and Mitchell, E.P., J. Appl. Cryst. (1996) 29, 584- 587.
d e s c r i p t i o n
Order Information
Each Crystal Screen Cryo kit contains 50 unique reagents. To order individual reagents, use
Custom Shop

catalog number listed below. Refer to page 36 for further details.

Cat. No. Name Description Price
HR2-122 Crystal Screen Cryo 10 ml, tube format $285.00
HR2-914-** Crystal Screen Cryo Custom Shop 185 ml $138.00
** = reagent number 1-50
C R Y S T A L S C R E E N C R Y O

Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com 22
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The protein crystal above is a LFA-1 I-domain/Inhibitor
complex. Crystallization conditions obtained using
Crystal Screen Cryo from Hampton Research.
Courtesy of Joerg Kallen.
Novartis Pharma AG
s u c c e s s s t o r y
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1. 0.02 M Calcium chloride dihydrate, 0.1 M Sodium acetate trihydrate pH 4.6,
30% v/v (+
/
-)-2-Methyl-2,4-pentanediol
2. 0.26 M Potassium sodium tartrate tetrahydrate, 35% v/v Glycerol
3. 0.26 M Ammonium phosphate monobasic, 35% v/v Glycerol
4. 0.075 M Tris hydrochloride pH 8.5, 1.5 M Ammonium sulfate, 25% v/v Glycerol
5. 0.2 M Sodium citrate tribasic dihydrate, 0.1 M HEPES sodium pH 7.5,
30% v/v (+
/
-)-2-Methyl-2,4-pentanediol
6. 0.16 M Magnesium chloride hexahydrate, 0.08 M Tris hydrochloride pH 8.5,
24% w/v Polyethylene glycol 4,000, 20% v/v Glycerol
7. 0.07 M Sodium cacodylate trihydrate pH 6.5, 0.98 M Sodium acetate trihydrate, 30% v/v Glycerol
8. 0.14 M Sodium citrate tribasic dihydrate, 0.07 M Sodium cacodylate trihydrate pH 6.5,
21% v/v 2-Propanol, 30% v/v Glycerol
9. 0.17 M Ammonium acetate, 0.085 M Sodium citrate tribasic dihydrate pH 5.6,
25.5% w/v Polyethylene glycol 4,000, 15% v/v Glycerol
10. 0.17 M Ammonium acetate, 0.085 M Sodium acetate trihydrate pH 4.6,
25.5% w/v Polyethylene glycol 4,000, 15% v/v Glycerol
11. 0.07 M Sodium citrate tribasic dihydrate pH 5.6, 0.7 M Ammonium phosphate monobasic,
30% v/v Glycerol
12. 0.18 M Magnesium chloride hexahydrate, 0.09 M HEPES sodium pH 7.5, 27% v/v 2-Propanol,
10% v/v Glycerol
13. 0.2 M Sodium citrate tribasic dihydrate, 0.1 M Tris hydrochloride pH 8.5,
30% v/v Polyethylene glycol 400
14. 0.19 M Calcium chloride dihydrate, 0.095 M HEPES sodium pH 7.5, 26.6% v/v Polyethylene glycol 400,
5% v/v Glycerol
15. 0.17 M Ammonium sulfate, 0.085 M Sodium cacodylate trihydrate pH 6.5,
25.5% w/v Polyethylene glycol 8,000, 15% v/v Glycerol
16. 0.075 M HEPES sodium pH 7.5, 1.125 M Lithium sulfate monohydrate, 25% v/v Glycerol
17. 0.17 M Lithium sulfate monohydrate, 0.085 M Tris hydrochloride pH 8.5,
25.5% w/v Polyethylene glycol 4,000, 15% v/v Glycerol
18. 0.16 M Magnesium acetate tetrahydrate, 0.08 M Sodium cacodylate trihydrate pH 6.5,
16% w/v Polyethylene glycol 8,000, 20% v/v Glycerol
19. 0.16 M Ammonium acetate, 0.08 M Tris hydrochloride pH 8.5, 24% v/v 2-Propanol, 20% v/v Glycerol
20. 0.16 M Ammonium sulfate, 0.08 M Sodium acetate trihydrate pH 4.6,
20% w/v Polyethylene glycol 4,000, 20% v/v Glycerol
21. 0.2 M Magnesium acetate tetrahydrate, 0.1 M Sodium cacodylate trihydrate pH 6.5,
30% v/v (+
/
-)-2-Methyl-2,4-pentanediol
22. 0.17 M Sodium acetate trihydrate, 0.085 M Tris hydrochloride pH 8.5,
25.5% w/v Polyethylene glycol 4,000, 15% v/v Glycerol
23. 0.2 M Magnesium chloride hexahydrate, 0.1 M HEPES sodium pH 7.5, 30% v/v Polyethylene glycol 400
24. 0.14 M Calcium chloride dihydrate, 0.07 M Sodium acetate trihydrate pH 4.6, 14% v/v 2-Propanol,
30% v/v Glycerol
25. 0.07 M Imidazole pH 6.5, 0.7 M Sodium acetate trihydrate, 30% v/v Glycerol
26. 0.2 M Ammonium acetate, 0.1 M Sodium citrate tribasic dihydrate pH 5.6,
30% v/v (+
/
-)-2-Methyl-2,4-pentanediol
27. 0.14 M Sodium citrate tribasic dihydrate, 0.07 M HEPES sodium pH 7.5, 14% v/v 2-Propanol,
30% v/v Glycerol
28. 0.17 M Sodium acetate trihydrate, 0.085 M Sodium cacodylate trihydrate pH 6.5,
25.5% w/v Polyethylene glycol 8,000, 15% v/v Glycerol
29. 0.065 M HEPES sodium pH 7.5, 0.52 M Potassium sodium tartrate tetrahydrate, 35% v/v Glycerol
30. 0.17 M Ammonium sulfate, 25.5% w/v Polyethylene glycol 8,000, 15% v/v Glycerol
31. 0.17 M Ammonium sulfate, 25.5% w/v Polyethylene glycol 4,000, 15% v/v Glycerol
32. 1.5 M Ammonium sulfate, 25% v/v Glycerol
33. 3.6 M Sodium formate, 10% v/v Glycerol
34. 0.07 M Sodium acetate trihydrate pH 4.6, 1.4 M Sodium formate, 30% v/v Glycerol
35. 0.075 M HEPES sodium pH 7.5, 0.6 M Sodium phosphate monobasic monohydrate,
0.6 M Potassium phosphate monobasic, 25% v/v Glycerol
36. 0.065 M Tris hydrochloride pH 8.5, 5.2% w/v Polyethylene glycol 8,000, 35% v/v Glycerol
37. 0.07 M Sodium acetate trihydrate pH 4.6, 5.6% w/v Polyethylene glycol 4,000, 30% v/v Glycerol
38. 0.09 M HEPES sodium pH 7.5, 1.26 M Sodium citrate tribasic dihydrate, 10% v/v Glycerol
39. 0.085 M HEPES sodium pH 7.5, 1.7% v/v Polyethylene glycol 400, 1.7 M Ammonium sulfate,
15% v/v Glycerol
40. 0.095 M Sodium citrate tribasic dihydrate pH 5.6, 19% v/v 2-Propanol,
19% w/v Polyethylene glycol 4,000, 5% v/v Glycerol
41. 0.085 M HEPES sodium pH 7.5, 8.5% v/v 2-Propanol, 17% w/v Polyethylene glycol 4,000,
15% v/v Glycerol
42. 0.04 M Potassium phosphate monobasic, 16% w/v Polyethylene glycol 8,000,
20% v/v Glycerol
43. 24% w/v Polyethylene glycol 1,500, 20% v/v Glycerol
44. 0.1 M Magnesium formate dihydrate, 50% v/v Glycerol
45. 0.16 M Zinc acetate dihydrate, 0.08 M Sodium cacodylate trihydrate pH 6.5,
14.4% w/v Polyethylene glycol 8,000, 20% v/v Glycerol
46. 0.16 M Calcium acetate hydrate, 0.08 M Sodium cacodylate trihydrate pH 6.5,
14.4% w/v Polyethylene glycol 8,000, 20% v/v Glycerol
47. 0.08 M Sodium acetate trihydrate pH 4.6, 1.6 M Ammonium sulfate, 20% v/v Glycerol
48. 0.08 M Tris hydrochloride pH 8.5, 1.6 M Ammonium phosphate monobasic, 20% v/v Glycerol
49. 0.8 M Lithium sulfate monohydrate, 1.6% w/v Polyethylene glycol 8,000, 20% v/v Glycerol
50. 0.4 M Lithium sulfate monohydrate, 12% w/v Polyethylene glycol 8,000, 20% v/v Glycerol
C R Y S TA L S C R E E N C R Y O F O R M U L AT I O N
Range from 4.0 to 9.0
p
H
o r g a n i c
p o l y m e r
n o n - v o l a t i l e
o r g a n i c
s a l t
(+/-)-2-Methyl-2,4-pentanediol
2-Propanol
Ammonium acetate
Ammonium phosphate
Ammonium sulfate
Calcium acetate
Calcium chloride
Lithium sulfate
Magnesium acetate
Magnesium chloride
Magnesium formate
Potassium phosphate
K/Na tartrate
Sodium acetate
Sodium citrate
Sodium formate
Sodium phosphate
Zinc acetate
Polyethylene glycol 400
Polyethylene glycol 1,500
Polyethylene glycol 4,000
Polyethylene glycol 8,000
b u f f e r
HEPES sodium
Imidazole
Sodium acetate
Sodium cacodylate
Sodium citrate
TRIS hydrochloride
Glycerol
c r y o p r o c t e c a n t
crystal screen cryo
f a c t o r s
Crystal of RNA tetraloop.
Courtesy of Imre Berger and Li Su.
MIT
s u c c e s s s t o r y
n
Crystallization screen for nucleic acid
fragments
n
Screen a matrix of pH, polyamine, & ions
n
Can be used with ribozymes, pseudoknots,
RNA, & DNA
n
Uses MPD as the primary precipitant
a p p l i c a t i o n
f e a t u r e s
The Nucleic Acid Mini Screen is an efficient
screen formulated to assist in the determi-
nation of preliminary crystallization condi-
tions of nucleic acid fragments.
1
The for-
mulation is based upon the publication, A
Highly Efficient 24 Condition Matrix for the
Crystallization of Nucleic Acid Fragments
where the preliminary crystallization condi-
tions of 35 nucleic acids were determined
utilizing this formulation.
1
Samples include
DNA-Drug complexes, C-Tetrad and G-Quartet Motifs, RNA oligomers, and others. By using 1 to 4 mM
oligonucleotide stock concentration, the screen requires less than 100 l of sample. The screen is typically
performed at 4C although room temperature incubations can also be performed. To evaluate the effect of
equilibration kinetics as well as initial and final sample concentrations, the screens are typically performed
using the hanging or sitting drop vapor diffusion method with two drops on a slide (1 l + 2 l and 2 l +
2 l), side by side. The 24 well format of the mini screen provides for a fast setup and uses small amounts of
sample. This makes it possible to cost-effectively screen many sequences as well as variations of a particular
sequence. The composition of the Nucleic Acid Mini Screen allows one to apply the formulation to other
nucleic acids such as deoxy- and ribozymes, pseudoknots, and tRNAs.
Each Nucleic Acid Mini Screen kit contains 24 unique reagents, 1.0 ml each plus a 250 ml volume of dehy-
drant (35% v/v MPD). All solutions are sterile filtered and formulated with ultra-pure Type 1 water.
References
1. Berger, et al., A Highly Effective 24 Condition Matrix for the Crystallization of Nucleic Acid Fragments. Acta Cryst. Section D.
(1996) Vol. D52 Part 3, 465-468.
2. Adams, A., Nucleic Acids Research (2002) Vol. 30, 719-725.
3. Crystallization and preliminary X-ray diffraction analysis of an Escherichia coli tRNAGly acceptor-stem microhelix. C. Frster, M. Perbandt,
A. B. E. Brauer, S. Brode, J. P. Frste, C. Betzel and V. A. Erdmann. Acta Cryst. (2007). F63, 46-48.
d e s c r i p t i o n
Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com 24
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N U C L E I C A C I D M I N I S C R E E N

Order Information
Each Nucleic Acid Mini Screen kit contains 24 unique reagents plus dehydrant.
Cat. No. Name Description Price
HR2-118 Nucleic Acid Mini Screen 1 ml, tube format + 250 ml bottle $195.00
5.5 to 7.0
using
40 mM Sodium
cacodylate
pH
35% v/v MPD
Lithium
Sodium
Potassium
Magnesium
Strontium
Barium
chlorides
Cobalt hexamine
Spermine tetra-HCl
polyamines
+
+
2
+
+
2
dehydrant
Tube # Precipitant
1. 10% v/v (+/-)-2-Methyl-2,4-pentanediol
2. 10% v/v (+/-)-2-Methyl-2,4-pentanediol
3. 10% v/v (+/-)-2-Methyl-2,4-pentanediol
4. 10% v/v (+/-)-2-Methyl-2,4-pentanediol
5. 10% v/v (+/-)-2-Methyl-2,4-pentanediol
6. 10% v/v (+/-)-2-Methyl-2,4-pentanediol
7. 10% v/v (+/-)-2-Methyl-2,4-pentanediol
8. 10% v/v (+/-)-2-Methyl-2,4-pentanediol
9. 10% v/v (+/-)-2-Methyl-2,4-pentanediol
10. 10% v/v (+/-)-2-Methyl-2,4-pentanediol
11. 10% v/v (+/-)-2-Methyl-2,4-pentanediol
12. 10% v/v (+/-)-2-Methyl-2,4-pentanediol
13. 10% v/v (+/-)-2-Methyl-2,4-pentanediol
14. 10% v/v (+/-)-2-Methyl-2,4-pentanediol
15. 10% v/v (+/-)-2-Methyl-2,4-pentanediol
16. 10% v/v (+/-)-2-Methyl-2,4-pentanediol
17. 10% v/v (+/-)-2-Methyl-2,4-pentanediol
18. 10% v/v (+/-)-2-Methyl-2,4-pentanediol
19. 10% v/v (+/-)-2-Methyl-2,4-pentanediol
20. 10% v/v (+/-)-2-Methyl-2,4-pentanediol
21. 10% v/v (+/-)-2-Methyl-2,4-pentanediol
22. 10% v/v (+/-)-2-Methyl-2,4-pentanediol
23. 10% v/v (+/-)-2-Methyl-2,4-pentanediol
24. 10% v/v (+/-)-2-Methyl-2,4-pentanediol
Tube # Buffer
1. 40 mM Sodium cacodylate pH 5.5
2. 40 mM Sodium cacodylate pH 5.5
3. 40 mM Sodium cacodylate pH 5.5
4. 40 mM Sodium cacodylate pH 5.5
5. 40 mM Sodium cacodylate pH 6.0
6. 40 mM Sodium cacodylate pH 6.0
7. 40 mM Sodium cacodylate pH 6.0
8. 40 mM Sodium cacodylate pH 6.0
9. 40 mM Sodium cacodylate pH 6.0
10. 40 mM Sodium cacodylate pH 6.0
11. 40 mM Sodium cacodylate pH 6.0
12. 40 mM Sodium cacodylate pH 6.0
13. 40 mM Sodium cacodylate pH 6.0
14. 40 mM Sodium cacodylate pH 7.0
15. 40 mM Sodium cacodylate pH 7.0
16. 40 mM Sodium cacodylate pH 7.0
17. 40 mM Sodium cacodylate pH 7.0
18. 40 mM Sodium cacodylate pH 7.0
19. 40 mM Sodium cacodylate pH 7.0
20. 40 mM Sodium cacodylate pH 7.0
21. 40 mM Sodium cacodylate pH 7.0
22. 40 mM Sodium cacodylate pH 7.0
23. 40 mM Sodium cacodylate pH 7.0
24. 40 mM Sodium cacodylate pH 7.0
Tube # Polyamine
1. 20 mM Cobalt hexamine
2. 20 mM Cobalt hexamine
3. 20 mM Cobalt hexamine
4. 20 mM Cobalt hexamine
5. 12 mM Spermine tetrahydrochloride
6. 12 mM Spermine tetrahydrochloride
7. 12 mM Spermine tetrahydrochloride
8. 12 mM Spermine tetrahydrochloride
9. 12 mM Spermine tetrahydrochloride
10. 12 mM Spermine tetrahydrochloride
11. 12 mM Spermine tetrahydrochloride
12. 12 mM Spermine tetrahydrochloride
13. 12 mM Spermine tetrahydrochloride
14. 12 mM Spermine tetrahydrochloride
15. 12 mM Spermine tetrahydrochloride
16. 12 mM Spermine tetrahydrochloride
17. 12 mM Spermine tetrahydrochloride
18. 12 mM Spermine tetrahydrochloride
19. 12 mM Spermine tetrahydrochloride
20. 12 mM Spermine tetrahydrochloride
21. 12 mM Spermine tetrahydrochloride
22. 12 mM Spermine tetrahydrochloride
23. 12 mM Spermine tetrahydrochloride
24. 12 mM Spermine tetrahydrochloride
Tube # Monovalent Ion
1. None
2. 80 mM Sodium chloride
3. 12 mM Sodium chloride,
80 mM Potassium chloride
4. 40 mM Lithium chloride
5. 80 mM Potassium chloride
6. 80 mM Potassium chloride
7. 80 mM Sodium chloride
8. 80 mM Sodium chloride
9. 80 mM Sodium chloride,
12 mM Potassium chloride
10. 12 mM Sodium chloride,
80 mM Potassium chloride
11. 80 mM Sodium chloride
12. 80 mM Potassium chloride
13. None
14. 80 mM Potassium chloride
15. 80 mM Potassium chloride
16. 80 mM Sodium chloride
17. 80 mM Sodium chloride
18. 80 mM Sodium chloride,
12 mM Potassium chloride
19. 12 mM Sodium chloride,
80 mM Potassium chloride
20. 80 mM Sodium chloride
21. 80 mM Potassium chloride
22. 40 mM Lithium chloride
23. 40 mM Lithium chloride
24. None
Tube # Divalent Ion
1. 20 mM Magnesium chloride
2. 20 mM Magnesium chloride
3. None
4. 20 mM Magnesium chloride
5. 20 mM Magnesium chloride
6. None
7. 20 mM Magnesium chloride
8. None
9. 20 mM Magnesium chloride
10. None
11. 20 mM Barium chloride
12. 20 mM Barium chloride
13. 80 mM Strontium chloride
14. 20 mM Magnesium chloride
15. None
16. 20 mM Magnesium chloride
17. None
18. 20 mM Magnesium chloride
19. None
20. 20 mM Barium chloride
21. 20 mM Barium chloride
22. 80 mM Strontium chloride,
20 mM Magnesium chloride
23. 80 mM Strontium chloride
24. 80 mM Strontium chloride,
20 mM Magnesium chloride
N U C L E I C A C I D M I N I S C R E E N F O R M U L A T I O N
Crystal is that of the intact canine lymphoma
antibody. Preliminary crystallization conditions
determined using the Low Ionic Strength Screen.
Courtesy of Lisa Harris.
University of California, Irvine
s u c c e s s s t o r y
n
Crystallization screen for intact monoclonal
antibodies, monoclonal antibody fragments,
& proteins less soluble at low ionic strength
n
Screen a complete pH profile (2-12) at low
ionic strength
n
Enhanced temperature effects due to low
ionic strength
n
Monodisperse PEG 3350
a p p l i c a t i o n
f e a t u r e s
The Low Ionic Strength Screen is effective for
determining the preliminary crystallization
conditions of intact monoclonal antibodies
1
.
However, this screen is not just an intact
antibody screen. The screen has effectively
determined the preliminary crystallization
conditions for numerous monoclonal anti-
body fragments as well as other soluble
proteins. The screen should be utilized as a
low ionic strength crystallization screen for
proteins where this strategy could be effective in determining preliminary crystallization screens.
In this screen, the concentration of a high purity, monodisperse PEG 3,350 is varied from 4 to 24% w/v (4, 8,
12, 16, 20 and 24% w/v) versus a pH range of 2 to 12 (2, 3, 3.5, 4, 4.5, 5, 5.5, 6.0, 6.5, 7, 7.5, 8, 8.5, 9, 9.5, 10,
11, 12). Stock buffer concentrations are 50 mM. Final buffer concentration in the drop is typically 10 mM.
Unique features of this screen: (1) Low ionic strength. Buffer concentration is supplied as 50 mM, result-
ing in an initial drop concentration of 10 mM (Drop = 4 l sample, 2 l buffer, 5 l precipitant); (2)
The Polyethylene glycol (PEG) 3,350 is a special, high-purity, monodisperse preparation with an Mr of
3300-3400. Most PEGs of this molecular weight have an Mr of plus or minus 500 rather than 50; (3) An
extremely broad range of pH 2 to 12 is sampled. At low ionic strength, the effects of pH and temperature
upon sample solubility are amplified. Hence this screen allows one to critically evaluate the effects of tem-
perature and pH upon sample solubility and crystallization. It is recommended the screen be repeated at
several temperatures between 4C and 37C to take advantage of the low ionic strength feature.
The format of this screen is different than other screens offered by Hampton Research. The screen is sup-
plied as a set of eighteen 50 mM buffers, six PEG 3,350 concentrations, and a single dehydrant, PEG 3,350.
This formulation allows one to screen up to 108 conditions if desired. The formulation is designed to offer
options. Coarse sample of pH and/or precipitant concentration can be performed with 24 or even 12 drops.
This will eliminate large regions of sampling space and conserve sample. Subsequent screens and perhaps
even optimization can encompass an increasingly finer grid matrix of pH, precipitant concentration, as well
as other variables significant for sample crystallization. The protocol requires the following pipetting steps
for a typical vapor diffusion experiment: (1) Pipet dehydrant to reservoir; (2) Pipet drop; (3) Pipet buffer
to drop; (4) Pipet precipitant to drop.
Each kit contains 24 unique reagents of varying pH and PEG 3,350 concentration. Buffers of 50 mM are sup-
plied for pH 2, 3, 3.5, 4, 4.5, 5, 5.5, 6, 6.5, 7, 7.5, 8, 8.5, 9, 9.5, 10, 11, and 12 in 1 ml volumes. The precipitant
Polyethylene glycol 3,350 is supplied as 4, 8, 12, 16, 20, and 24% w/v in 1 ml volumes. Dehydrant is supplied
as 250 ml of 24% PEG 3,350. Combining each pH and precipitant supplied can generate 108 crystallization
screening conditions. All solutions are sterile filtered and formulated with ultra-pure Type 1 water.
References
1. Harris, et al., Crystallization of intact monoclonal antibodies, Proteins: Structure, Function, and Genetics (1995) 23:285-289.
Order Information
Each Low Ionic Strength Screen kit contains 24 unique reagents plus dehydrant.
Cat. No. Name Description Price
HR2-120 Low Ionic Strength Screen 1 ml, tube format + 250 ml bottle $195.00
d e s c r i p t i o n
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l o w I o n I C s T r e n G T H s C r e e n

Precipitant
4% w/v Polyethylene glycol 3,350
8% w/v Polyethylene glycol 3,350
12% w/v Polyethylene glycol 3,350
16% w/v Polyethylene glycol 3,350
20% w/v Polyethylene glycol 3,350
24% w/v Polyethylene glycol 3,350
Buffer
0.05 M Potassium chloride pH 2.0
0.05 M Citric acid pH 3.0
0.05 M Citric acid pH 3.5
0.05 M Citric acid pH 4.0
0.05 M Citric acid pH 4.5
0.05 M Citric acid pH 5.0
0.05 M Citric acid pH 5.5
0.05 M MES monohydrate pH 6.0
0.05 M BIS-TRIS pH 6.5
0.05 M Imidazole pH 7.0
0.05 M HEPES pH 7.5
0.05 M Tris pH 8.0
0.05 M Tris pH 8.5
0.05 M Glycine pH 9.0
0.05 M Glycine pH 9.5
0.05 M Glycine pH 10.0
0.05 M Sodium phosphate dibasic pH 11.0
0.05 M Sodium phosphate dibasic pH 12.0
Dehydrant
24% w/v Polyethylene glycol 3,350
Low Ionic Strength
Screen Components
n
Primary, secondary or orthoganol crystallization
screen for biological macromolecules
n
For use with soluble proteins and membrane
proteins
n
A methodology to uncover different crystal
forms
n
Optimization screening in conjunction with
preliminary crystallization conditions
n
ThermoFluor

assays
n
Solubility, folding and stability studies
n
Developed at Hampton Research
n
Screens a portfolio of small molecules for their
ability to establish stabilizing, intermolecular,
hydrogen bonding, hydrophobic and electro-
static interactions which could promote lattice
formation and crystallization

n
Organic salts & acids

n
Biologically active small molecules

n
Amino acids and peptide

n
Macromolecular digests
n
Modular screen design allows Silver Bullets to
be screened against virtually any crystallization
reagent
n
Original & Bio Formulations
n
A combinatorial library of more than 1,090
chemicals, of which more than 400 are unique
n
Tube or Deep Well block format
a p p l i c a t i o n s
f e a t u r e s
An alternative approach to macromolecular crystallization.
The Silver Bullets formulation and methodology screens
validated small molecule libraries against a few select crys-
tallization reagents as an effective, orthogonal approach
to macromolecular crystallization. The formulation offers
an alternative and efficient approach that is an excellent
complement to current crystallization method-ologies.
Silver Bullets is a library of small molecules that have been
shown to promote crystal lattice formation. X-ray diffraction analysis has demonstrated the reagents have
the ability to:
Stabilize the conformation of the protein
Perturb the interaction of the protein with the solvent
Participate in forming important lattice contacts
Build the crystal lattice by forming reversible cross-links between the macromolecules in the crystal
Results with the Silver Bullets approach have been very encouraging, with more than twice as many
proteins being crystallized overall as were crystallized from controls free of any small molecules.
1-3
There
have been frequent occasions where some exceptional result has been obtained for specific proteins,
and numerous examples where new or unusual crystal forms were obtained. X-ray diffraction analysis has
revealed the small molecules in the crystal lattice, involved at the centers of hydrogen bonding networks
and electrostatic interaction.
3
Silver Bullets can be used as a primary crystallization screen; as a secondary or orthogonal screen to pro-
duce crystals when traditional screens are not successful; as an optimization screen in conjunction with
preliminary crystallization conditions; as a methodology to uncover different crystal forms; to promote
intramolecular interactions that may stabilize a macromolecules conformation and promote crystallization.
Silver Bullets solutions are designed for use with the Silver Bullets PEG/Tacsimate Crystallization Reagents
but may also be used with other crystallization reagents.
The Silver Bullets kits are composed of 96 solutions in either screw cap tubes or a single Deep Well block
(Greiner 786261) HT format. Each reagent is a mixture of small molecules or macromolecular digest in
0.02 M HEPES sodium pH 6.8 buffer. Each Silver Bullets solution is supplied in a 0.25 ml volume. Each
solution contains between 2 and 20 small molecules. Silver Bullets reagents include: Organic salts and acids
Biologically active molecules Amino acids and peptides Macromolecular digests. Silver Bullets Bio
reagents include: Small organic molecules, organic salts, and organic acids Biologically active molecules,
co-factors, and ligands Amino acids, peptides, nucleotides, drugs, and carbohydrates Biochemical
pathway intermediates.
References
1. Searching for silver bullets: An alternative strategy for crystallizing macromolecules. Alexander McPherson and Bob Cudney. Journal of Structural Biology
156 (2006) 387-406.
2. A novel strategy for the crystallization of proteins: X-ray diffraction validation. Steven B. Larson, John S. Day, Robert Cudney, and Alexander McPherson.
Acta Cryst. (2007) D63, 310-318.
3. Development of an alternative approach to protein crystallization. McPherson, Alexander; Nguyen, Chieniang; Larson, Steven B; Day, John S; Cudney,
Bob. J Struct Funct Genomics, Volume 8, Number 4, December 2007, 193-198.
d e s c r i p t i o n
Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com 26
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S I L V E R B U L L E T S

s I l v e r b u l l e T s b I o

Mellitic acid, a Silver Bullet for Trypsin

Order Information
Silver Bullets and Silver Bullets Bio kits each contain 96 unique reagents. To order individual
reagents, use Custom Shop

catalog numbers listed below. Refer to page 36 for further details.

Cat. No. Name Description Price
HR2-078 Silver Bullets 0.25 ml, tube format $490.00
HR2-096 Silver Bullets HT 0.25 ml, Deep Well block format $490.00
HR2-080 Silver Bullets Bio 0.25 ml, tube format $490.00
HR2-088 Silver Bullets Bio HT 0.25 ml, Deep Well block format $490.00
HR2-090 PEG/Tacsimate pH 5.8 1 ml, Deep Well block format $185.00
HR2-092 PEG/Tacsimate pH 6.8 1 ml, Deep Well block format $185.00
HR2-094 PEG/Tacsimate pH 7.8 1 ml, Deep Well block format $185.00
HR2-996-** Silver Bullets Custom Shop 0.25 ml $65.00
HR2-988-** Silver Bullets Bio Custom Shop 0.25 ml $65.00

** = reagent number 1-96
27
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organi c salts
organi c aci ds
compati ble wi th a di verse
array of reagents
f a c t o r s
silver bullets

macromolecular di gests
ami no aci ds
pepti des
combi natori al
formulati on
combi natori al
formulati on
bi ologi cally acti ve small molecules
ami no aci ds
pepti des
nucleoti des
carbohydrates
small organi c molecules
organi c salts
organi c aci ds
compati ble wi th a di verse
array of reagents
bi ologi cally acti ve molecules co- factors li gands
f a c t o r s
engage in multiple, strong hydrogen bonds within the protein
form hydrophobic interactions with the protein
become involved in electrostatic interactions with the protein
enhance, disrupt and alter the pattern of clustering in the critical
nucleus of the crystal
develop stabilizing lattice interactions within the crystal
serve as transient stabilizing intermediaries
c o m p o s e d o f c o m p o u n d s w i t h t h e a b i l i t y t o
Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com 28
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Manipulate sample-sample & sample-solvent
interactions to enhance or alter sample
solubility
n
18 classes of reagents
n
Highly concentrated (10x) reagent formulation
n
96 unique reagents
n
Crystallization or sample solubility
optimization
n
Tube or Deep Well block format
a p p l i c a t i o n
f e a t u r e s
Additive Screen is a library of small molecules
that can affect the solubility and crystalliza-
tion of biological macromolecules, including
both soluble and membrane proteins. These
small molecules can perturb and manipulate
sample-sample and sample-solvent interac-
tions, as well as perturb water structure
which can alter and improve both the solubil-
ity and crystallization of a sample. Additives
can stabilize or engender conformity by spe-
cific interaction with the macromolecules. There are numerous reports of the use of additives to improve
the quality and size of macromolecular crystals.
1-5

Additive Screen contains 96 unique reagents, 1 ml each, tube format.
Additive Screen HT contains 96 unique reagents, 1 ml each, in a single Deep Well block format.
References
1. Crystallization of membrane proteins., Edited by Hartmut Michel, CRC Press (1991).
2. Crystallization of Nucleic Acids and Proteins: A Practical Approach., Edited by A. Ducruix and R. Giege, Oxford University Press (1992).
3. Cudney, R., et al., Screening and optimization strategies for macromolecular crystal growth., Acta Cryst. (1994) D50, 414-423.
4. Sousa R., Use of glycerol, polyols and other protein structure stabilizing agents in protein crystallization., Acta Cryst. (1995) D51, 271-277.
5. Trakhanov, S. and Quiocho, F.A., Influence of divalent cations on protein crystallization., Protein Science (1995) 4, 9, 1914-1919.
Order Information
Each Additive Screen kit contains 96 unique reagents. To order individual reagents, use Custom
Shop

catalog number listed below. Refer to page 36 for further details.

Cat. No. Name Description Price
HR2-428 Additive Screen 1 ml, tube format $490.00
HR2-138 Additive Screen HT 1 ml, Deep Well block format $490.00
HR2-428-** Additive Screen Custom Shop 1 ml $65.00
** = reagent number 1-96
d e s c r i p t i o n
Initial crystals grown using Crystal Screen Reagent
Number 9. Crystals were approximately 100 microns
in size. With the addition of Dextran Sulfate from the
Additive Screen kit, larger crystals were produced as
pictured above (approximately 250 microns).
Courtesy of Ranjit K. Deka.
University of Texas Southwestern Medical Center
s u c c e s s s t o r y
A D D I T I V E S C R E E N

a d d I T I v e s C r e e n H T

Tube # Additive
73. (G1) 30% w/v Trimethylamine N-oxide dihydrate
74. (G2) 30% w/v 1,6-Hexanediol
75. (G3) 30% v/v (+/-)-2-Methyl-2,4-pentanediol
76. (G4) 50% v/v Polyethylene glycol 400
77. (G5) 50% v/v Jeffamine M-600

pH 7.0
78. (G6) 40% v/v 2,5-Hexanediol
79. (G7) 40% v/v ()-1,3-Butanediol
80. (G8) 40% v/v Polypropylene glycol P 400
81. (G9) 30% v/v 1,4-Dioxane
82. (G10) 30% v/v Ethanol
83. (G11) 30% v/v 2-Propanol
84. (G12) 30% v/v Methanol
85. (H1) 40% v/v 1,4-Butanediol
86. (H2) 40% v/v tert-Butanol
87. (H3) 40% v/v 1,3-Propanediol
88. (H4) 40% v/v Acetonitrile
89. (H5) 40% v/v Formamide
90. (H6) 40% v/v 1-Propanol
91. (H7) 5% v/v Ethyl acetate
92. (H8) 40% v/v Acetone
93. (H9) 0.25% v/v Dichloromethane
94. (H10) 7% v/v 1-Butanol
95. (H11) 40% v/v 2,2,2-Trifluoroethanol
96. (H12) 40% v/v 1,1,1,3,3,3-Hexafluoro-2-propanol
A D D I T I V E S C R E E N F O R M U L A T I O N
Tube # Additive
1. (A1) 0.1 M Barium chloride dihydrate
2. (A2) 0.1 M Cadmium chloride hydrate
3. (A3) 0.1 M Calcium chloride dihydrate
4. (A4) 0.1 M Cobalt(II) chloride hexahydrate
5. (A5) 0.1 M Copper(II) chloride dihydrate
6. (A6) 0.1 M Magnesium chloride hexahydrate
7. (A7) 0.1 M Manganese(II) chloride tetrahydrate
8. (A8) 0.1 M Strontium chloride hexahydrate
9. (A9) 0.1 M Yttrium(II) chloride hexahydrate
10. (A10) 0.1 M Zinc chloride
11. (A11) 0.1 M Iron(III) chloride hexahydrate
12. (A12) 0.1 M Nickel(II) chloride hexahydrate
13. (B1) 0.1 M Chromium(III) chloride hexahydrate
14. (B2) 0.1 M Praseodymium(III) acetate hydrate
15. (B3) 1.0 M Ammonium sulfate
16. (B4) 1.0 M Potassium chloride
17. (B5) 1.0 M Lithium chloride
18. (B6) 2.0 M Sodium chloride
19. (B7) 0.5 M Sodium fluoride
20. (B8) 1.0 M Sodium iodide
21. (B9) 2.0 M Sodium thiocyanate
22. (B10) 1.0 M Potassium sodium tartrate tetrahydrate
23. (B11) 1.0 M Sodium citrate tribasic dihydrate
24. (B12) 1.0 M Cesium chloride
Tube # Additive
25. (C1) 1.0 M Sodium malonate pH 7.0
26. (C2) 0.1 M L-Proline
27. (C3) 0.1 M Phenol
28. (C4) 30% v/v Dimethyl sulfoxide
29. (C5) 0.1 M Sodium bromide
30. (C6) 30% w/v 6-Aminohexanoic acid
31. (C7) 30% w/v 1,5-Diaminopentane dihydrochloride
32. (C8) 30% w/v 1,6-Diaminohexane
33. (C9) 30% w/v 1,8-Diaminooctane
34. (C10) 1.0 M Glycine
35. (C11) 0.3 M Glycyl-glycyl-glycine
36. (C12) 0.1 M Taurine
37. (D1) 0.1 M Betaine hydrochloride
38. (D2) 0.1 M Spermidine
39. (D3) 0.1 M Spermine tetrahydrochloride
40. (D4) 0.1 M Hexammine cobalt(III) chloride
41. (D5) 0.1 M Sarcosine
42. (D6) 0.1 M Trimethylamine hydrochloride
43. (D7) 1.0 M Guanidine hydrochloride
44. (D8) 0.1 M Urea
45. (D9) 0.1 M b-Nicotinamide adenine dinucleotide
hyd.
46. (D10) 0.1 M Adenosine-5'-triphosphate disodium salt
47. (D11) 0.1 M TCEP hydrochloride
48. (D12) 0.01 M GSH (L-Glutathione reduced),
0.01 M GSSG (L-Glutathione oxidized)
Tube # Additive
49. (E1) 0.1 M EDTA sodium salt
50. (E2) 5% w/v Polyvinylpyrrolidone K15
51. (E3) 30% w/v Dextran sulfate sodium salt (Mr 5,000)
52. (E4) 40% v/v Pentaerythritol ethoxylate (3/4 EO/OH)
53. (E5) 10% w/v Polyethylene glycol 3,350
54. (E6) 30% w/v D(+)-Glucose monohydrate
55. (E7) 30% w/v Sucrose
56. (E8) 30% w/v Xylitol
57. (E9) 30% w/v D-Sorbitol
58. (E10) 12% w/v myo-Inositol
59. (E11) 30% w/v D-(+)-Trehalose dihydrate
60. (E12) 30% w/v D-(+)-Galactose
61. (F1) 30% v/v Ethylene glycol
62. (F2) 30% v/v Glycerol
63. (F3) 3.0 M NDSB-195
64. (F4) 2.0 M NDSB-201
65. (F5) 2.0 M NDSB-211
66. (F6) 2.0 M NDSB-221
67. (F7) 1.0 M NDSB-256
68. (F8) 0.15 mM CYMAL

-7
69. (F9) 20% w/v Benzamidine hydrochloride hydrate
70. (F10) 5% w/v n-dodecyl-N,N-dimethylamine-N-oxide
71. (F11) 5% w/v n-Octyl-b-D-glucoside
72. (F12) 5% w/v n-Dodecyl-b-D-maltoside
Concentrations are actual formulations.
29
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a p p l i c a t i o n
f e a t u r e s
Detergent Screen kits are designed to allow the
rapid and convenient evaluation of 96 unique
detergent reagents for their ability to influence
the solubility and crystallization of the sample.
These screens are designed to be compatible
with most popular crystallization reagents.
The detergents in these kits are capable of
manipulating hydrophobic sample-sample inter-
actions which can lead to non-specific aggre-
gation, and prevent or interfere with sample
crystallization. The detergents also perturb water
structure which may play a role in sample crystal-
lization.
Non-specific aggregation is a common deterrent
to the crystallization of soluble macromolecules
as well as membrane proteins. There is exten-
sive literature demonstrating the effectiveness of including detergents in the crystallization trial towards
preventing non-specific aggregation due to hydrophobic interactions and hence improving crystallization.
1
The kits allow one to screen the most effective detergents used in crystal growth.
The Detergent Screen kits are preformulated so that simple pipetting is all that is required to screen the
detergents with the hanging or sitting drop vapor diffusion technique. Screens with these kits are usually
performed after preliminary crystallization conditions have been determined, although ab initio screens
are also practical. The kits are recommended for both soluble and membrane proteins where non-specific
aggregation is a suspected complication or where one simply wishes to screen detergents as an optimiza-
tion variable. The screens are suitable for hanging drop, sitting drop, microbatch, free interface diffusion
and sandwich drop crystallization methodologies.
Each Detergent Screen kit contains 96 unique reagents. 0.25 ml of detergent solution is preformulated at
10 times the reported CMC (unless otherwise noted) in sterile filtered water.
References
1. Crystallization of membrane proteins. Edited by Hartmut Michel, CRC Press, (1991).
Order Information
Each Detergent Screen kit contains 96 unique reagents. To order individual reagents, use Custom Shop

catalog number listed below. Refer to page 36 for further details.
Cat. No. Name Description Price
HR2-408 Detergent Screen 0.25 ml, tube format $490.00
HR2-406 Detergent Screen HT 0.25 ml, Deep Well block format $490.00
HR2-406-** Detergent Screen Custom Shop 0.5 ml $65.00
** = reagent number 1-96
d e s c r i p t i o n
n
Prevent and manipulate non-specific
aggregation due to hydrophobic interactions
n
Kits include popular detergents used in the
crystallization of membrane proteins

n
Ionic detergents

n
Non-ionic detergents

n
Zwitterionic detergents

n
Non-detergent Sulfobetaines

n
Synthetic lipids
n
Useful with soluble proteins where
hydrophobic interactions limit sample solubility
n
Detergents are compatible with microbatch
under oil crystallization, as well as vapor
diffusion, free interface diffusion, & dialysis
D E T E R G E N T S C R E E N

d e T e r G e n T s C r e e n H T

Detergent molecule interacting with beta-lactamase.

Crystal above is that of -Lactamase. Crystal was
grown using 15% w/v PEG 6000, 50 mM HEPES
pH 7.0, and in the presence of CYMAL-5 and
CYMAL-6 detergent from the Hampton Research
Detergent Screen kit 1.
Courtesy of James Knox.
University of Connecticut,
Department of Molecular and Cell Biology
s u c c e s s s t o r y
D E T E R G E N T S C R E E N F O R M U L A T I O N
Well # Detergent CMC (mM)
1. (A1)
2. (A2)
3. (A3)
4. (A4)
5. (A5)
6. (A6)
7. (A7)
8. (A8)
9. (A9)
10. (A10)
11. (A11)
12. (A12)
13. (B1)
14. (B2)
15. (B3)
16. (B4)
17. (B5)
18. (B6)
19. (B7)
20. (B8)
21. (B9)
22. (B10)
23. (B11)
24. (B12)
BAM
n-Dodecyl-b-iminodipropionic acid,
monosodium salt
Dodecyltrimethylammonium chloride
CTAB
Deoxycholic acid, sodium salt
Sodium dodecyl sulfate
Sodium cholate
Sodium dodecanoyl sarcosine
ANAPOE

-X-305
IPTG
n-Hexadecyl-b-D-maltoside
ANAPOE

-58
n-Tetradecyl-b-D-maltoside
ANAPOE

-80
n-Tridecyl-b-D-maltoside
ANAPOE

-C12E9
ANAPOE

-20
Thesit

ANAPOE

-35
ANAPOE

-C13E8
ANAPOE

-C12E8
n-Dodecyl-b-D-maltoside
CYMAL

-7
ANAPOE

-X-114
None
None
0.046
1.0
6.0
8.27
14.0
14.4
None
None
0.0006
0.004
0.010
0.012
0.033
0.05
0.059
0.09
0.091
0.10
0.11
0.17
0.19
0.20
Well # Detergent CMC (mM)
25. (C1)
26. (C2)
27. (C3)
28. (C4)
29. (C5)
30. (C6)
31. (C7)
32. (C8)
33. (C9)
34. (C10)
35. (C11)
36. (C12)
37. (D1)
38. (D2)
39. (D3)
40. (D4)
41. (D5)
42. (D6)
43. (D7)
44. (D8)
45. (D9)
46. (D10)
47. (D11)
48. (D12)
ANAPOE

-C12E10
Sucrose monolaurate
CYMAL

-6
n-Undecyl-b-D-maltoside
ANAPOE

-X-405
TRITON

X-100
ANAPOE

-C10E6
n-Decyl-b-D-thiomaltoside
Octyl maltoside, fluorinated
ANAPOE

-C10E9
Big CHAP, deoxy
n-Decyl-b-D-maltoside
LDAO
n-Decanoylsucrose
n-Nonyl-b-D-thioglucoside
n-Nonyl-b-D-thiomaltoside
CYMAL

-5
n-Nonyl-b-D-maltoside
n-Nonyl-b-D-glucoside
HEGA

-10
MEGA -10
C8E5
CYMAL

-4
C8E4
0.20
0.30
0.56
0.59
0.81
0.90
0.90
0.90
1.02
1.3
1.4
1.8
2.0
2.5
2.9
3.2
5.0
6.0
6.5
7.0
7.0
7.1
7.6
8.0
Well # Detergent CMC (mM)
73. (G1)
74. (G2)
75. (G3)
76. (G4)
77. (G5)
78. (G6)
79. (G7)
80. (G8)
81. (G9)
82. (G10)
83. (G11)
84. (G12)
85. (H1)
86. (H2)
87. (H3)
88. (H4)
89. (H5)
90. (H6)
91. (H7)
92. (H8)
93. (H9)
94. (H10)
95. (H11)
96. (H12)
NDSB-195
NDSB-201
NDSB-211
NDSB-221
NDSB-256
ZWITTERGENT

3-14
n-Dodecyl-N,N-dimethylglycine
FOS-Choline

-12
FOS-Choline

-8, fluorinated
n-Undecyl-N,N-Dimethlamine-oxide
ZWITTERGENT

3-12
DDMAB
FOS-MEA

-10
CHAPS
CHAPSO
FOS-Choline

-10
n-Decyl-N,N-dimethylglycine
FOS-Choline

-9
ZWITTERGENT

3-10
CYCLOFOS
TM
-3
FOS-Choline

-8
ZWITTERGENT

3-08
LysoFos
TM
Choline 12
LysoFos
TM
Choline 10
N/A
N/A
N/A
N/A
N/A
0.40
1.5
1.5
2.2
3.21
4.0
4.3
5.25
8.0
8.0
11.0
19.0
39.5
40.0
43.0
114.0
330.0
0.7
7.0
Well # Detergent CMC (mM)
49. (E1)
50. (E2)
51. (E3)
52. (E4)
53. (E5)
54. (E6)
55. (E7)
56. (E8)
57. (E9)
58. (E10)
59. (E11)
60. (E12)
61. (F1)
62. (F2)
63. (F3)
64. (F4)
65. (F5)
66. (F6)
67. (F7)
68. (F8)
69. (F9)
70. (F10)
71. (F11)
72. (F12)
n-Octyl-b-D-thiomaltoside
n-Octyl-b-D-thioglucoside
Hexaethylene glycol monooctyl ether
DDAO
C-HEGA

-11
Pluronic

F-68
HECAMEG

n-Octyl-b-D-glucoside
n-Octanoylsucrose
MEGA-9
2,6-Dimethyl-4-heptyl-b-D-malto-pyranoside
n-Heptyl-b-D-thioglucopyranoside
n-Octyl-b-D-galactopyranoside
CYMAL

-3
C-HEGA

-10
HEGA

-9
Dimethyloctylphosphine oxide
MEGA-8
C-HEGA

-9
HEGA

-8
CYMAL

-2
n-Hexyl-b-D-glucopyranoside
C-HEGA

-8
CYMAL

-1
8.5
9.0
10.0
10.4
11.5
17.9
19.5
20.0
24.4
25.0
27.5
29.0
29.5
34.5
35.0
39.0
40.0
79.0
108.0
109.0
120.0
250.0
277.0
340.0
Iridium Compounds
H
1.008 1
Li
6.940 3
Na
22.99 11
K
39.09 19
Ca
40.08 20
Sc
45.10 21
Ti
47.90 22
V
50.95 23
Cr
52.01 24
Mn
54.93 25
Fe
55.84 26
Co
58.94 27
Ni
58.69 28
Cu
63.57 29
Zn
65.38 30
Ga
69.72 31
Ge
72.60 32
As
74.91 33
Se
78.96 34
Br
79.916 35
Kr
83.70 36
Mg
24.32 12
Be
9.02 4
Ar
39.944 18
Cl
35.457 17
S
32.06 16
P
30.98 15
Si
28.06 14
Al
26.97 13
B
10.82 5
C
12.01 6
N
14.008 7
O
16.00 8
F
19.00 9
Ne
20.183 10
He
4.003 2
Rb
85.48 37
Sr
87.63 38
Y
88.92 39
Zr
91.22 40
Nb
92.91 41
Mo
95.95 42
Tc
97.907 43
Ru
101.70 44
Rh
102.91 45
Pd
106.70 46
Ag
107.88 47
Cd
112.41 48
In
114.76 49
Sn
118.70 50
Sb
121.76 51
Te
127.61 52
I
126.92 53
Xe
131.30 54
Cs
132.91 55
Ba
137.36 56
La
138.92 57
Hf
178.60 72
Ta
180.88 73
W
183.92 74
Re
186.31 75
Os
190.20 76
Ir
193.10 77
Pt
195.23 78
Au
197.20 79
Hg
200.61 80
Tl
204.39 81
Pb
207.21 82
Bi
209.00 83
Po
208.98 84
At
209.99 85
Rn
222.02 86
Fr
223.02 87
Ra
226.03 88
Ac
227.03 89 90
to
103
58
to
71
Lanthanides
Actinides
Ce
140.13 58
Pr
140.92 59
Nd
144.27 60
Pm
144.91 61
Sm
150.43 62
Eu
152.00 63
Gd
156.90 64
Tb
159.20 65
Dy
162.46 66
Ho
164.94 67
Er
167.20 68
Tm
169.40 69
Yb
173.04 70
Lu
175.00 71
Th
232.12 90
Pa
231.04 91
U
238.07 92
Np
237.05 93
Pu
244.06 94
Am
243.06 95
Cm
247.07 96
Bk
247.07 97
Cf
242.06 98
Es
252.08 99
Fm
257.10 100
Md
258.10 101
No
259.10 102
Lr
260.11 103
Osmium Compounds
Silver Compounds
Platinum Compounds
Mercury Compounds Thallium Compounds
Lead Compounds Gold Compounds
Cadmium Compounds
Tungsten Compounds
Samarium Compounds
Lanthanum Compounds
Europium Compounds Gadolinium Compounds
Ytterbium Compounds
Dysprosium Compounds
Praseodymium Compounds
Neodymium Compounds Holmium Compounds
Rhenium Compounds
Lutetium Compounds
Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com 30
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Heavy atoms for multiple isomorphous
replacement
n
Convenient sets of popular heavy atoms
n
Each kit includes a heavy atom tutorial and
formulation guide
a p p l i c a t i o n
f e a t u r e s
A Heavy Atom Screen kit provides 50 mg
of each heavy atom in an o-ring screw cap
micro tube. This convenient format provides
sufficient material for the preparation of
numerous (15 x 100 mM or 1,500 x 1 mM)
small volume (100 l) fresh stock solutions
of heavy atoms. The quantity is sufficient for
screening and derivatization but avoids the
problem of storing large quantities of heavy
atoms. Individual heavy atoms from each
kit are available in larger quantities on a special order basis for researchers requiring larger amounts of
material.
References
1. Heavy-atom derivatization. Elspeth Garman and James W. Murray, Acta Cryst. (2003). D59, 1903-1913.
2. Towards a rational approach for heavy-atom derivative screening in protein crystallography. Johnson Agniswamy, M. Gordon Joyce, Carl H. Hammer,
and Peter D. Sun. Acta Cryst. (2008) D64, 354-367.
3. Screening for phasing atoms in protein crystallography. Titus J Boggon and Lawrence Shapiro. Structure 2000, Vol 8 No 7, 143-149.
d e s c r i p t i o n
H e a v y a T o M s C r e e n s
Protein crystal with the heavy atom derivative
Potassium osmate.
Kimberly J. Skinner, Structural Biology and Biophysics,
Pfizer Global R&D, Sandwich, Kent, United Kingdom.
s u c c e s s s t o r y
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Order Information
Each Heavy Atom Screen Pt kit contains 12 unique reagents. To order individual
reagents, use Custom Shop

catalog number listed below. Refer to page 36 for

further details.
Cat. No. Name Description Price
HR2-442 Heavy Atom Screen Pt 50 mg, tube format $418.00
HR2-442-** Heavy Atom Screen Pt 100 mg $80.00
Custom Shop
** = reagent number 1-12
Order Information
Each Heavy Atom Screen Au kit contains 6 unique reagents. To order individual
reagents, use Custom Shop

catalog number listed below. Refer to page 36 for

further details.
Cat. No. Name Description Price
HR2-444 Heavy Atom Screen Au 50 mg, tube format $275.00
HR2-444-** Heavy Atom Screen Au 100 mg $80.00
Custom Shop
** = reagent number 1-6
Order Information
Each Heavy Atom Screen Hg kit contains 15 unique reagents. To order individual
reagents, use Custom Shop

catalog number listed below. Refer to page 36 for

further details.
Cat. No. Name Description Price
HR2-446 Heavy Atom Screen Hg 50 mg, tube format $314.00
HR2-446-** Heavy Atom Screen Hg 100 mg $80.00
Custom Shop
** = reagent number 1-15
Heavy Atom Screen Pt (Platinum)
1. Potassium tetrachloroplatinate(II)
2. Ammonium tetrachloroplatinate(II)
3. Potassium hexachloroplatinate(IV)
4. Potassium tetranitroplatinate(II)
5. Potassium tetracyanoplatinate(II) hydrate
6. Dichloro(ethylenediamine)platinum(II)
7. Diammino Platinum Dinitrite
8. Potassium tetrabromoplatinate(II)
9. Potassium hexabromoplatinate(IV)
10. Platinum potassium iodide
11. Platinum potassium thiocyanate
12. Di--iodobis(ethylenediamine)diplatinum(II) nitrate
Heavy Atom Screen Au (Gold)
1. Gold(I) potassium cyanide
2. Potassium tetrachloroaurate(III) hydrate
3. Sodium tetrachloroaurate(III) dihydrate
4. Gold(III) chloride
5. Hydrogen tetrachloroaurate(III) trihydrate
6. Potassium tetrabromoaurate(III) dihydrate
Heavy Atom Screen Hg (Mercury)
1. Mersalyl acid
2. Ethyl Mercuric Phosphate
3. Mercury(II) chloride
4. Mercury(II) acetate
5. Ethylmercurithiosalicylic acid, sodium salt
6. Phenylmercury acetate
7. Mercury(II) potassium iodide
8. p-Chloromercuribenzoic acid
9. Ethylmercury chloride
10. Mercury(II) bromide
11. Mercury(II) iodide
12. Mercury(II) nitrate monohydrate
13. Mercury(II) cyanide
14. Mercury(II) oxide, yellow
15. Tetrakis(acetoxymercuri)methane
Heavy Atom Screen M2
1. Sodium tungstate dihydrate
2. Ammonium tetrathiotungstate(VI)
3. Samarium(III) chloride hexahydrate
4. Samarium(III) acetate hydrate
5. Samarium(III) nitrate hexahydrate
6. Lanthanum(III) nitrate hexahydrate
7. Europium(III) nitrate hexahydrate
8. Europium(III) chloride hexahydrate
9. Gadolinium(III) chloride hydrate
10. Lutetium(III) chloride hexahydrate
11. Lutetium(III) acetate hydrate
12. Ytterbium(III) chloride hydrate
13. Dysprosium(III) chloride hexahydrate
14. Praseodymium(III) chloride heptahydrate
15. Neodymium(III) chloride hydrate
16. Holmium(III) chloride hexahydrate
17. Potassium hexachlororhenate(IV)
18. Potassium perrhenate
Order Information
Each Heavy Atom Screen M1 kit contains 19 unique reagents. To order individual
reagents, use Custom Shop

catalog number listed below. Refer to page 36 for

further details.
Cat. No. Name Description Price
HR2-448 Heavy Atom Screen M1 50 mg, tube format $343.00
HR2-448-** Heavy Atom Screen M1 100 mg $80.00
Custom Shop
** = reagent number 1-19
Order Information
Each Heavy Atom Screen M2 kit contains 18 unique reagents. To order individual
reagents, use Custom Shop

catalog number listed below. Refer to page 36 for

further details.
Cat. No. Name Description Price
HR2-450 Heavy Atom Screen M2 50 mg, tube format $320.00
HR2-450-** Heavy Atom Screen M2 100 mg $80.00
Custom Shop
** = reagent number 1-18
Heavy Atom Screen M1
1. Thallium(III) chloride hydrate
2. Thallium(I) chloride
3. Thallium(III) acetate hydrate
4. Lead(II) acetate trihydrate
5. Lead(II) nitrate
6. Lead(II) chloride
7. Silver nitrate
8. Cadmium chloride hydrate
9. Cadmium iodide
10. Potassium hexachloroiridate(IV)
11. Iridium(III) chloride hydrate
12. Sodium hexachloroiridate(III) hydrate
13. Ammonium hexachloroiridate(III) hydrate
14. Potassium hexanitroiridium(III)
15. Potassium osmate(VI) dihydrate
16. Ammonium hexabromoosmate(IV)
17. Potassium hexachloroosmate(IV)
18. Osmium(III) chloride hydrate
19. Acetoxytrimethyllead(IV)
Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com 32
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Produce heavy atom derivative of biological
macromolecules for phasing
n
I3C kit for soaking or cocrystallization
n
Phasing using SAD or SIRAS
n
Ready-to-use, no weighing required
a p p l i c a t i o n
f e a t u r e s
I3C can be used for heavy atom derivatization
of biological macromolecules for subsequent
single wavelength anomalous dispersion
(SAD) or single isomorphous replacement
plus anomalous scattering (SIRAS).
The two carboxylic acid groups and one
amino group of I3C can interact by way of
hydrogen bonds with both the backbone and
side chains of proteins. This can result in a
relatively highoccupancy of the bound ligand
I3C. The three iodine atoms per I3C mol-
ecule provide for a strong anomalous signal.
The three iodine atoms in I3C form an equilateral triangle with 6.0 side lengths. These triangular struc-
tures can readily be identified in the anomalous electron density map.
Kit Contents
12 aliquots of I3C (280 mg each) (5-Amino-2,4,6-triiodoisophthalic acid CAS number 35453-19-1)
12 aliquots of 2.0 M Lithium hydroxide (650 l each)
User Guide
References
1. A magic triangle for experimental phasing of macromolecules. Beck et al. Acta Cryst. D64, 1179, 2008.
2. Structure determination of the cancer-associated Mycoplasma hyorhinis protein Mhp37. Sippel et al. Acta Cryst. D64, 1172, 2008.
3. 5-Amino-2,4,6 triiodo-isophthalic acid monohydrate. Beck et al. Acta Cryst. E64:1286, 2008.
Order Information
Cat. No. Name Description Price
HR3-133 I3C Phasing Kit 24 tubes $155.00
d e s c r i p t i o n
1 3 C P H a s I n G k I T

Crystal of cytochrome P450 Cyp121 from Mycobacterium tuberculosis with red color due
to the ferric heme iron. Crystal diffracts to 1.4Ang at Proxima1/Soleil, France.
Pascal Belin, iBiTec-s/CEA, Saclay, France.
Crystal of human phosphatase.
Chris Gray at the Institute of Cancer Research,
London, United Kingdom.
Crystal Image.
Allan DArcy, Switzerland.
Crystals of apo kinase.
Initial hit from Hampton Research Grid Screen Salt HT.
Michelle Quiles and Annie Hassell,
GlaxoSmithKline, RTP, North Carolina, USA.
c u s t o m s h o p
c r y s t a l l i z a t i o n
r e a g e n t s
Protein crystal grown with additive.
Ranjit Deka, University of Texas Southwest Medical Center, USA.
T A B L E O F C O N T E N T S
36 - 37 c u s t o m s h o p

c r y s t a l l i z a t i o n r e a g e n t s
P A G E S
C R Y S T A L S C R E E N L I T E

Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com
The Hampton Research Custom Shop delivers ready-to-use crystallization
reagents for screening, optimizing and producing crystals. The reagents are
formulated at the time of order, using the same production protocol, chemi-
cals, and quality control used when making the original Hampton Research
crystallization kit.
Formulating the Custom Shop reagents at the time of order ensures consis-
tency as well as saves time and offers convenience. They are shipped within 48
hours. Custom Shop reagents may not be returned for refund or exchange.
To order Custom Shop reagents, use the catalog numbers listed below. If you
have any questions, please feel free to contact Customer Service.
When reproducing and optimizing reagent conditions, please be sure to also
consider the Hampton Research Optimize and StockOptions reagents
and kits.
d e s c r i p t i o n
Heavy Atom Pt (Platinum) - 100 mg $80.00
Catalog Number: HR2-442-**
** = reagent number 1-12
For example, reagent number 1 = HR2-442-01
Additive Screen - 1.0 ml $65.00
Catalog Number: HR2-428-**
** = reagent number 1-96
For example, reagent number A1 = HR2-428-01
Crystal Screen - 185 ml $138.00
Catalog Number: HR2-910-**
** = reagent number 1-50
For example, reagent number 1 = HR2-910-01
Crystal Screen Cryo - 185 ml $138.00
Catalog Number: HR2-914-**
** = reagent number 1-50
For example, reagent number 1 = HR2-914-01
Crystal Screen Lite - 185 ml $138.00
Catalog Number: HR2-916-**
** = reagent number 1-50
For example, reagent number 1 = HR2-916-01
Grid Screen PEG/LiCl - 185 ml $138.00
Catalog Number: HR2-928-**
** = reagent number A1-D6
For example, reagent number A1 = HR2-928-A1
Grid Screen Sodium Chloride - 185 ml $138.00
Catalog Number: HR2-932-**
** = reagent number A1-D6
For example, reagent number A1 = HR2-932-A1
Grid Screen Sodium Malonate - 185 ml $138.00
Catalog Number: HR2-947-**
** = reagent number A1-D6
For example, reagent number A1 = HR2-947-A1
Crystal Screen 2 - 185 ml $138.00
Catalog Number: HR2-912-**
** = reagent number 1-48
For example, reagent number 1 = HR2-912-01
Heavy Atom Au (Gold) - 100 mg $80.00
Catalog Number: HR2-444-**
** = reagent number 1-6
For example, reagent number 1 = HR2-444-01
Heavy Atom Hg (Mercury) - 100 mg $80.00
Catalog Number: HR2-446-**
** = reagent number 1-16
For example, reagent number 1 = HR2-446-01
Heavy Atom M1 - 100 mg $80.00
Catalog Number: HR2-448-**
** = reagent number 1-19
For example, reagent number 1 = HR2-448-01
Heavy Atom M2 - 100 mg $80.00
Catalog Number: HR2-450-**
** = reagent number 1-18
For example, reagent number 1 = HR2-450-01
Index - 185 ml $138.00
Catalog Number: HR2-944-**
** = reagent number 1-96
For example, reagent number 1 = HR2-944-01
Quik Screen - 185 ml $138.00
Catalog Number: HR2-921-**
** = reagent number A1-D6
For example, reagent number A1 = HR2-921-A1
MembFac - 185 ml $138.00
Catalog Number: HR2-920-**
** = reagent number 1-48
For example, reagent number 1 = HR2-920-01
Natrix - 185 ml $138.00
Catalog Number: HR2-918-**
** = reagent number 1-48
For example, reagent number 1 = HR2-918-01
PCT - 185 ml $138.00
Catalog Number: HR2-940-**
** = reagent number A1-B2
For example, reagent number A1 = HR2-940-A1
PEG/Ion Screen - 185 ml $138.00
Catalog Number: HR2-922-**
** = reagent number 1-48
For example, reagent number 1 = HR2-922-01
PEG/Ion Screen 2 - 185 ml $138.00
Catalog Number: HR2-998-**
** = reagent number 1-48
For example, reagent number 1 = HR2-998-01
PEGRx 1 - 185 ml $138.00
Catalog Number: HR2-982-**
** = reagent number 1-48
For example, reagent number 1 = HR2-982-01
PEGRx 2 - 185 ml $138.00
Catalog Number: HR2-984-**
** = reagent number 1-48
For example, reagent number 1 = HR2-984-01
SaltRx 1 - 185 ml $138.00
Catalog Number: HR2-907-**
** = reagent number 1-48
For example, reagent number 1 = HR2-907-01
SaltRx 2 - 185 ml $138.00
Catalog Number: HR2-909-**
** = reagent number 1-48
For example, reagent number 1 = HR2-909-01
Silver Bullets - 0.25 ml $65.00
Catalog Number: HR2-996-**
** = reagent number 1-96
For example, reagent number A1 = HR2-996-01
Silver Bullets Bio - 0.25 ml $65.00
Catalog Number: HR2-988-**
** = reagent number 1-96
For example, reagent number A1 = HR2-988-01
Detergent Screen - 0.5 ml $65.00
Catalog Number: HR2-406-**
** = reagent number 1-96
For example, reagent number A1 = HR2-406-01
Grid Screen MPD - 185 ml $138.00
Catalog Number: HR2-930-**
** = reagent number A1-D6
For example, reagent number A1 = HR2-930-A1
Grid Screen PEG 6000 - 185 ml $138.00
Catalog Number: HR2-926-**
** = reagent number A1-D6
For example, reagent number A1 = HR2-926-A1
C u s T o M s H o P

C R Y S T A L L I Z A T I O N R E A G E N T S
Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com 36
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Grid Screen Ammonium Sulfate - 185 ml $138.00
Catalog Number: HR2-924-**
** = reagent number A1-D6
For example, reagent number A1 = HR2-924-A1
StockOptions ADA - 185 ml $208.00
Catalog Number: HR2-997-**
** = reagent number 1-13
For example, reagent number 1 = HR2-998-01
StockOptions Imidazole - 185 ml $162.00
Catalog Number: HR2-995-**
** = reagent number 1-17
For example, reagent number 1 = HR2-995-01
StockOptions MES - 185 ml $138.00
Catalog Number: HR2-943-**
** = reagent number 1-20
For example, reagent number 1 = HR2-943-01
StockOptions pH - 185 ml
Catalog Number: HR2-941-**
Reagents 1-39 $139.00
Reagents 40-45 $151.00
** = reagent number 1-45
For example, reagent number 1 = HR2-941-01
StockOptions Sodium Acetate - 185 ml $150.00
Catalog Number: HR2-933-**
** = reagent number 1-21
For example, reagent number 1 = HR2-933-01
StockOptions Sodium Cacodylate - 185 ml $221.00
Catalog Number: HR2-939-**
** = reagent number 1-24
For example, reagent number 1 = HR2-939-01
StockOptions Sodium Citrate - 185 ml $173.00
Catalog Number: HR2-935-**
** = reagent number 1-24
For example, reagent number 1 = HR2-935-01
StockOptions Sodium HEPES - 185 ml $190.00
Catalog Number: HR2-931-**
** = reagent number 1-15
For example, reagent number 1 = HR2-931-01
StockOptions Tris - 185 ml $169.00
Catalog Number: HR2-900-**
** = reagent number 1-21
For example, reagent number 1 = HR2-900-01
StockOptions Tris Hydrochloride - 185 ml $139.00
Catalog Number: HR2-937-**
** = reagent number 1-21
For example, reagent number 1 = HR2-937-01
StockOptions Bicine - 185 ml $139.00
Catalog Number: HR2-999-**
** = reagent number 1-15
For example, reagent number 1 = HR2-999-01
StockOptions Bis-Tris - 185 ml $151.00
Catalog Number: HR2-906-**
** = reagent number 1-21
For example, reagent number 1 = HR2-906-01
StockOptions Bis-Tris Propane - 185 ml $201.00
Catalog Number: HR2-993-**
** = reagent number 1-33
For example, reagent number 1 = HR2-993-01
StockOptions Citric Acid - 185 ml $144.00
Catalog Number: HR2-904-**
** = reagent number 1-44
For example, reagent number 1 = HR2-904-01
StockOptions HEPES - 185 ml $172.00
Catalog Number: HR2-902-**
** = reagent number 1-15
For example, reagent number 1 = HR2-902-01
37
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Crystals growing on a bubble.

Michelle Dechene, North Carolina State University, North Carolina, USA.
Here is a recipe to try:
Mosaicity is about 0.5
t0 0.6
Reagent:
Crystal Screen Cryo
Reagent 23
Mix equal amounts of
Glucose Isomerase
and reagent. Vapor
diffusion method.

Mount crystal in
CryoLoop.

Mosaicity may be
a bit more in this
reagent and the
unit cell will shrink a
o p t i m i z e

c r y s t a l l i z a t i o n g r a d e
r e a g e n t s
Protein crystal.
Alexey Rak, Max-Planck-Institut fur Molekulare Physiologie, Department of Physical Biochemistry, Dortmund, Germany.
P r e - C r y s t a l l i z a t i o n T e s t
I n d e x
S a l t R x
P E G / I o n S c r e e n
C r y s t a l S c r e e n
C r y s t a l S c r e e n 2
C r y s t a l S c r e e n L i t e
C r y s t a l S c r e e n C r y o
G r i d S c r e e n s
q u i k s c r e e n & q u i k o p t i m i z e
L o w I o n i c S t r e n g t h S c r e e n
N u c l e i c A c i d M i n i S c r e e n
N a t r i x
M e m b f a c
S t o c k o p t i o n s S c r e e n s
A d d i t i v e S c r e e n
D e t e r g e n t S c r e e n
H e a v y a t o m s c r e e n s
C r y o p r o - C r y o p r o t e c t a n t
XX
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T A B L E O F C O N T E N T S
40 - 42 o p t i m i z e

- p o l y m e r s
42 o p t i m i z e

- o r g a n i c s ( v o l a t i l e )
42 o p t i m i z e

- o r g a n i c s ( n o n - v o l a t i l e )
43 - 49 o p t i m i z e

- s a l t s
50 - 52 o p t i m i z e

- b u f f e r s
52 o p t i m i z e

- s o l u b i l i z i n g a g e n t s ( n d s b )
53 o p t i m i z e

- r e d u c i n g a g e n t
54 o p t i m i z e

- c r y o p r o t e c t a n t s
55 - 56 o p t i m i z e

- o i l s
57 o p t i m i z e

- s i l i c a h y d r o g e l k i t
57 o p t i m i z e

- l m a g a r o s e
58 o p t i m i z e

- i z i t c r y s t a l d y e

P A G E S
n
Crystallization grade reagents for formulating
screens or for optimization
n
Quantitive formulation
n
Lot to lot consistency
n
Synergistic with Hampton Research
screens & kits
n
Quality and convenience
n
Sterile filtered
a p p l i c a t i o n
f e a t u r e s
Optimize reagents are quantitatively formulated
macromolecular crystallization grade reagents designed
specifically for the crystallization of proteins, peptides,
and nucleic acids. Each Optimize solution is formulated
using high purity salts, polymers, and buffers. Sterile
filtered Optimize reagents are formulated at convenient,
ready to use concentrations. Optimize reagents remove
the guesswork and make the process of reproducing
preliminary screening conditions and general optimiza-
tion faster, easier, and more convenient. When using
Optimize reagents, a scientist moves directly from
the screen to the optimization with no time wasted
searching for and formulating salts, buffers, and viscous
polymers. Each Optimize reagent is supplied with a
Certificate of Analysis as documentation of reagent qual-
ity and performance. Optimize buffer reagents are sup-
plied with calibrated titration curves to make titration
easy, fast, and accurate.
d e s c r i p t i o n
P O L Y M E R S
Ethylene imine polymer
HR2-599 50% solution 200 ml $78.00
Pentaerythritol ethoxylate (15/4 EO/OH)
HR2-745 50% v/v solution 200 ml $54.00
Pentaerythritol propoxylate (5/4 PO/OH)
HR2-739 50% v/v solution 200 ml $54.00
Synonyms: PEI or Poly(ethylene imine) or
Poly(ethyleneimine) solution or Ethyleneimine polymer solution
M
r
600,000 - 1,000,000 CAS [9002-98-6] EC No 205-793-9
Density: 1.08 g/mL at 20C Measured pH Range: 12.0 - 12.1 at 25C
Refractive Index n20/D: 1.450 Conductivity Range: 244.3 - 257.0 mS at 25C
Synonyms: Pentaerythritol ethoxylate (3/4 EO/OH)
C(CH2(OCH2CH2)nOH)4, N ~3.75 Average M
n
~797
CAS [30599-15-6] EC No 500-071-2
Density: 1.2 g/mL at 25C (lit.)
Measured pH Range: 5.2 - 6.0 at 25C
Refractive Index Range: 1.41401 - 1.41578 at 20C
Conductivity Range: 4.2 - 11.4 mS at 25C
Jeffamine

M-600

HR2-501 50% v/v solution pH 7.0 200 ml $94.00

HR2-503 100% solution (untitrated pH >12) 200 ml $94.00
Synonyms: O-(2-Aminopropyl)-O'-(2-methoxyethyl)polypropylene
glycol 500 or Polypropylene glycol 500 mono-2-aminoethyl
mono-2-methoxyethyl ether CH3OCH2CH2O[CH(CH3)CH2O]nCH2CH(NH2)
M
r
600 CAS [77110-54-4] Density: 0.983 g/mL at 20C
HR2-501: Refractive Index Range: 1.40754 - 1.40988 at 20C
Conductivity Range: 8.8 - 10.3 mS at 25C
HR2-503: Refractive Index Range: 1.44501 - 1.44546 at 20C
Conductivity Range: 0.0 - 0.1 mS at 25C
Synonyms: Pentaerythritol ethoxylate (15/4 EO/OH)
C(CH2(OCH2CH2)nOH)2 Average M
n
~270
CAS [30599-15-6] EC No 500-071-2
Density: 1.2 g/mL at 25C (lit.)
Measured pH Range: 7.5 - 8.7 at 25C
Refractive Index: 1.39719 at 20C
Conductivity Range: 37.8 - 40.5 mS at 25C
Pentaerythritol ethoxylate (3/4 EO/OH)
HR2-743 40% v/v solution 200 ml $46.00
Synonyms: Pentaerythritol propoxylate
C[CH2[OCH2CH(CH3)]nOH]4, N ~5 Average M
n
~426
CAS [9051-49-4] EC No 500-030-9
Density of 100% solution: 1.05 g/mL at 25C
Measured pH Range: 7.7 - 8.3 at 25C
Refractive Index: 1.40424 - 1.40433 at 20C
Conductivity Range: 3.8 - 4.7 mS at 25C
Synonyms: Pentaerythritol propoxylate
C(CH2(OCH2CH(CH3))nOH)4, N = 2.1 Average M
n
~629
CAS [9051-49-4] EC No 500-030-9
Density: 1.05 g/mL at 25C (lit.)
Measured pH Range: 4.3 - 4.6 at 25C
Refractive Index: 1.40418 at 20C
Conductivity Range: 6.1 - 6.9 mS at 25C
Synonyms: Poly(sodium acrylate) or Sodium polyacrylate
C3H3NaO2 M
r
94.04 CAS [9003-04-7]
Measured pH Range: 6.4 - 6.9 at 25C
Refractive Index Range: 1.40716 - 1.40821 at 20C Conductivity Range: 30.5 - 41.1 mS at 25C
Synonyms: PEG 200 H(OCH2CH2)nOH Mr 190 - 210
CAS [25322-68-3] EC No 500-038-2
Density: 1.124 g/mL at 20C
Measured pH Range: 8.0 - 10.2 at 25C
Refractive Index Range: 1.46008 - 1.46083 at 20C Conductivity Range: 2.5 - 2.8 mS at 25C
Pentaerythritol propoxylate (17/8 PO/OH)
HR2-741 50% v/v solution 200 ml $58.00
Poly(acrylic acid sodium salt) 5,100
HR2-773 50% w/v solution 200 ml $95.00
Polyethylene glycol 200
HR2-601 100% solution 200 ml $30.00
O P T I M I Z E

- R E A G E N T S
Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com 40
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Jeffamine

ED-2001 pH 7.0
HR2-597 50% w/v solution pH 7.0 200 ml $98.00
Synonyms: O,O'-Bis(2-aminopropyl) polypropylene
glycol-block-polyethylene glycol-block-polypropylene
glycol 1,900 CH3CH(NH2)CH2(OCH2CH2)nOCH2CH(NH2)CH3
M
r
~2,000 CAS [65605-36-9]
Refractive Index Range: 1.40637 - 1.40666 at 20C
Conductivity Range: 4.0 - 5.7 mS at 25C
pH before titration with Hydrochloric acid is > 10
Final measured pH of HR2-597 is 7.0
CH3 CH3 CH3
H2NCHCH2 NH2 (OCHCH2)a (OCH2CH2)b (OCH2CH)c
b 38.7, (a+c) 6.0
CH3 CH3 CH3
O O
x y z
O H2N NH2
( ( ( (((
y 39, (x+z) 6
41
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Polyethylene glycol 1,500
Polyethylene glycol 400
HR2-603 100% solution 200 ml $53.00
Polyethylene glycol 1,000
HR2-523 50% w/v solution 200 ml $90.00
Peroxides (as H2O2) 0.001%
Al ........................... 0.0005%
As ........................ 0.00001%
Ba .......................... 0.0005%
Bi ........................... 0.0005%
Ca ............................ 0.001%
Cd .......................... 0.0005%
Cl ............................. 0.005%
Co .......................... 0.0005%
Cr ........................... 0.0005%
Cu .......................... 0.0005%
Fe .......................... 0.0005%
K ................................ 0.02%
Li ............................ 0.0005%
Mg .......................... 0.0005%
Mn .......................... 0.0005%
Mo .......................... 0.0005%
Na .............................. 0.02%
Ni ........................... 0.0005%
Pb .......................... 0.0005%
SO4 .......................... 0.005%
Sr ........................... 0.0005%
Zn .......................... 0.0005%
HR2-525 50% w/v solution 200 ml $60.00
Polyethylene glycol 3,350 Monodisperse
HR2-591 flake 500 g $118.00
HR2-527 50% w/v solution 200 ml $80.00
HR2-837 25% w/v PEG 3,350, 0.1 M MES pH 5.8 100 ml $95.00
HR2-841 25% w/v PEG 3,350, 0.1 M HEPES Na pH 6.8 100 ml $95.00
HR2-845 25% w/v PEG 3,350, 0.1 M BIS-TRIS pH 7.8 100 ml $95.00
HR2-849 25% w/v PEG 3,350,
0.1 M BIS-TRIS propane pH 7.8 100 ml $95.00
P O L Y M E R S
Synonyms: PEG 400 H(OCH2CH2)nOH Mr 380 - 420
CAS [25322-68-3] EC No 500-038-2 Density: 1.126 g/mL at 20C
Measured pH Range: 9.1 - 9.7 at 25C
Refractive Index Range: 1.46686 - 1.46761 at 20C
Conductivity Range: 2.5 - 2.8 mS at 25C
Synonyms: PEG 1,000 H(OCH2CH2)nOH Mr 950 - 1050
CAS [25322-68-3] EC No 500-038-2
Measured pH Range: 6.5 - 7.7 at 25C
Refractive Index Range: 1.40078 - 1.40104 at 20C
Conductivity Range: 23.1 - 37.5 mS at 25C
Synonyms: PEG 3,350 H(OCH2CH2)nOH Mr 3,300 - 3,400
CAS [25322-68-3] EC No 500-038-2
HR2-527: Measured pH Range: 7.9 - 9.7 at 25C
Refractive Index Range: 1.40164 - 1.40208 at 20C
Conductivity Range: 44.0 - 56.5 mS at 25C
HR2-837: Measured Conductivity: 826 mS at 25C
HR2-841: Measured Refractive Index: 1.37193 at 20C
Measured Conductivity: 4.4 mS at 25C
HR2-845: Measured Conductivity: 379 mS at 25C
HR2-849: Measured Conductivity: 3.7 mS at 25C
Polyethylene glycol 4,000
HR2-605 flake 500 g $80.00
HR2-529 50% w/v solution 200 ml $80.00
Synonyms: PEG 4,000 H(OCH2CH2)nOH Mr 3,500 - 4,500
CAS [25322-68-3] EC No 500-038-2
HR2-529: Measured pH Range: 6.9 - 9.5 at 25C
Refractive Index Range: 1.40198 - 1.40279 at 20C
Conductivity Range: 38.6 - 46.9 mS at 25C
Polyethylene glycol monomethyl ether 550
HR2-611 100% solution 200 ml $85.00
Synonyms: Methoxypolyethylene glycol or mono-Methyl polyethylene glycol or PEG MME 550
CH3O(CH2CH2O)nH Mr 470 - 530 CAS [9004-74-4] EC No 215-801-2
Purity > 100.0% Density: 1.089 g/mL at 25C (lit.) Measured pH Range: 8.5 - 10.5 at 25C
Refractive Index Range: 1.46144 - 1.46187 at 20C Conductivity Range: 0.3 - 0.5 mS at 25C
Polyethylene glycol monomethyl ether 2,000
HR2-613 50% w/v solution 200 ml $85.00
Synonyms: Methoxypolyethylene glycol 2,000 or
mono-Methyl polyethylene glycol 2,000 or PEG MME 2,000
CH3O(CH2CH2O)nH CAS [9004-74-4] EC No 215-801-2
Purity > 95.0% Density: 1.094 g/mL at 25C (lit.) Measured pH Range: 7.0 - 8.8 at 25C
Refractive Index Range: 1.40135 - 1.40161 at 20C Conductivity Range: 28.0 - 60.2 mS at 25C
Synonyms: PEG 1,500 H(OCH
2CH2)nOH Mr 1,400 - 1,600
CAS [25322-68-3] EC No 500-038-2
Measured pH Range: 6.1 - 8.7 at 25C
Refractive Index Range: 1.40117 - 1.40127 at 20C
Conductivity Range: 39.7 - 48.5 mS at 25C
Polyethylene glycol 6,000
HR2-513 flake 500 g $122.00
HR2-533 50% w/v solution 200 ml $80.00
DNases .............. none detected
RNases .............. none detected
Proteases .......... none detected
Phosphatases .... none detected
Peroxides (H2O2) ..... 0.001%
Al ............................ 0.0005%
As ......................... 0.00001%
Ba ........................... 0.0005%
Bi ............................ 0.0005%
Ca ............................. 0.001%
Cd ........................... 0.0005%
Cl .............................. 0.005%
Co ........................... 0.0005%
Cr ............................ 0.0005%
Cu ........................... 0.0005%
Fe ........................... 0.0005%
K ................................. 0.02%
Li ............................. 0.0005%
Mg ........................... 0.0005%
Mn ........................... 0.0005%
Mo ........................... 0.0005%
Na ............................... 0.02%
Ni ............................ 0.0005%
Pb ........................... 0.0005%
SO
4
........................... 0.005%
Sr ............................ 0.0005%
Zn ........................... 0.0005%
Synonyms: PEG 6,000 H(OCH2CH2)nOH Mr 5,000 - 7,000
CAS [25322-68-3] EC No 500-038-2
HR2-533: Measured pH Range: 6.2 - 9.1 at 25C
Refractive Index Range: 1.40209 - 1.40249 at 20C
Conductivity Range: 32.5 - 39.6 mS at 25C
Polyethylene glycol 8,000
HR2-515 flake 500 g $122.00
HR2-535 50% w/v solution 200 ml $80.00
DNases ............. none detected
RNases ............. none detected
Proteases ......... none detected
Phosphatases ... none detected
Peroxides (as H2O2) 0.001%
Al ........................... 0.0005%
As ........................ 0.00001%
Ba .......................... 0.0005%
Bi ........................... 0.0005%
Ca ............................ 0.001%
Cd .......................... 0.0005%
Cl ............................. 0.005%
Co .......................... 0.0005%
Cr ........................... 0.0005%
Cu .......................... 0.0005%
Fe .......................... 0.0005%
K ................................ 0.02%
Li ............................ 0.0005%
Mg .......................... 0.0005%
Mn .......................... 0.0005%
Mo .......................... 0.0005%
Na .............................. 0.02%
Ni ........................... 0.0005%
Pb .......................... 0.0005%
SO
4 .......................... 0.005%
Sr ........................... 0.0005%
Zn .......................... 0.0005%
Synonyms: PEG 8,000 H(OCH2CH2)nOH Mr 7,000 - 9,000
CAS [25322-68-3] EC No 500-038-2
HR2-535: Measured pH Range: 6.6 - 9.5 at 25C
Refractive Index Range: 1.40206 - 1.40261 at 20C
Conductivity Range: 30.4 - 39.3 mS at 25C
Polyethylene glycol 10,000
HR2-607 50% w/v solution 200 ml $80.00
Synonyms: PEG 10,000 H(OCH2CH2)nOH Mr 8,500 - 11,500
CAS [25322-68-3] EC No 500-038-2
Measured pH Range: 6.4 - 8.6 at 25C
Refractive Index Range: 1.40165 - 1.40254 at 20C Conductivity Range: 39.4 - 48.3 mS at 25C
Polyethylene glycol 20,000
HR2-609 30% w/v solution 200 ml $80.00
Synonyms: PEG 20,000 H(OCH2CH2)nOH Average Mn ~16,000 - 24,000
CAS [25322-68-3] EC No 500-038-2
Measured pH Range: 4.6 - 10.2 at 25C
Refractive Index Range: 1.37438 - 1.37476 at 20C Conductivity Range: 45.3 - 64.5 mS at 25C
O
OH H
3
C
[ [
n
Polyethylene glycol 300
HR2-517 100% solution 200 ml $45.00
Synonyms: PEG 300 H(OCH2CH2)nOH Mr 285 - 315
CAS [25322-68-3] EC No 500-038-2 Density: 1.125 g/mL at 20C
Measured pH Range: 8.0 - 8.5 at 25C
Refractive Index Range: 1.46492 - 1.46535 at 20C
Conductivity Range: 7.7 - 8.1 mS at 25C
Peroxides (as H2O2) 0.001%
Al ........................... 0.0005%
As ........................ 0.00001%
Ba .......................... 0.0005%
Bi ........................... 0.0005%
Ca ............................ 0.001%
Cd .......................... 0.0005%
Cl ............................. 0.005%
Co .......................... 0.0005%
Cr ........................... 0.0005%
Cu .......................... 0.0005%
Fe .......................... 0.0005%
K .................................. 0.2%
Li ............................ 0.0005%
Mg .......................... 0.0005%
Mn .......................... 0.0005%
Mo .......................... 0.0005%
Na ................................ 0.2%
Ni ........................... 0.0005%
Pb .......................... 0.0005%
SO
4 .......................... 0.005%
Sr ........................... 0.0005%
Zn .......................... 0.0005%
O P T I M I Z E

- R E A G E N T S
Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com 42
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P O L Y M E R S
Polyethylene glycol monomethyl ether 5,000
HR2-615 50% w/v solution 200 ml $85.00
Polyvinylpyrrolidone K 15
HR2-769 50% w/v solution 200 ml $45.00
Polypropylene glycol P 400
HR2-771 100% solution 200 ml $30.00
1,4-Dioxane
HR2-617 100% solution 100 ml $48.00
2-Propanol
HR2-619 100% solution 200 ml $46.00
DNases................none detected
RNases................none detected
Proteases............none detected
Phosphatases......none detected
Al .......................... 0.00005%
Ba ......................... 0.00001%
Bi .......................... 0.00001%
Ca ......................... 0.00005%
Cd ....................... 0.000005%
Co ....................... 0.000002%
Cr ........................ 0.000002%
Cu ....................... 0.000002%
Fe ......................... 0.00001%
K ........................... 0.00005%
Li ........................... 0.00001%
Mg ......................... 0.00001%
Mn ....................... 0.000002%
Mo ......................... 0.00001%
Na ......................... 0.00005%
Ni ........................ 0.000002%
Pb ......................... 0.00001%
Sr .......................... 0.00001%
Zn ......................... 0.00001%
Al .......................... 0.00005%
Ba ......................... 0.00001%
Bi .......................... 0.00001%
Ca ......................... 0.00005%
Cd ....................... 0.000005%
Co ....................... 0.000002%
Cr ........................ 0.000002%
Cu ....................... 0.000002%
Fe ......................... 0.00001%
K ........................... 0.00005%
Li ........................... 0.00001%
Mg ......................... 0.00001%
Mn ....................... 0.000002%
Mo ......................... 0.00001%
Na ......................... 0.00005%
Ni ........................ 0.000002%
Pb ......................... 0.00001%
Sr .......................... 0.00001%
Zn ......................... 0.00001%
O R G A N I C S ( V O L A T I L E )
Ethylene glycol
HR2-621 100% solution 100 ml $34.00
A ........................ 0.00005%
Ba ...................... 0.00001%
Bi ....................... 0.00001%
Ca ...................... 0.00005%
Cd .................... 0.000005%
Cl ........................... 0.001%
Co .................... 0.000002%
Cr ..................... 0.000002%
Cu .................... 0.000002%
Fe ...................... 0.00005%
K ........................ 0.00005%
Li ........................ 0.00001%
Mg ...................... 0.00001%
Mn .................... 0.000005%
Mo ...................... 0.00001%
Na ........................ 0.0001%
Ni ..................... 0.000002%
Pb ...................... 0.00001%
SO4 ........................ 0.001%
Sr ....................... 0.00001%
Zn ...................... 0.00002%
Glycerol
HR2-623 100% solution 100 ml $61.00
DNases ........... None detected
RNases ........... None detected
Proteases ....... None detected
Phosphatases . None detected
Ag ........................ 0.0005%
Al ......................... 0.0001%
As ...................... 0.00001%
Ba ........................ 0.0001%
Bi ......................... 0.0001%
Ca ........................ 0.0005%
Cd ........................ 0.0001%
Cl ......................... 0.0001%
Co ........................ 0.0001%
Cr ......................... 0.0001%
Cu ....................... 0.0001%
Fe ........................ 0.0001%
K ............................ 0.002%
Li .......................... 0.0001%
Mg ........................ 0.0001%
Mn ........................ 0.0001%
Mo ........................ 0.0001%
NH4
+
.................... 0.0005%
Na .......................... 0.002%
Ni ......................... 0.0001%
Pb ........................ 0.0001%
SO4 ........................ 0.001%
Sr ......................... 0.0001%
Tl .......................... 0.0005%
Zn ........................ 0.0001%
1,6-Hexanediol
HR2-625 6.0 M (71% w/v) solution 200 ml $52.00
(+/-)-2-Methyl-2,4-pentandiol
HR2-627 100% solution 200 ml $62.00
O R G A N I C S ( N O N - V O L A T I L E )
Synonyms: Methoxypolyethylene glycol 5,000 or
mono-Methyl polyethylene glycol 5,000 or PEG MME 5,000
CH3O(CH2CH2O)nH CAS [9004-74-4] EC No 215-801-2
Purity > 95.0% Measured pH Range: 7.7 - 8.9 at 25C
Refractive Index Range: 1.40199 - 1.40236 at 20C Conductivity Range: 19.8 - 25.7 mS at 25C
Synonyms: 1,2-Ethanediol
C2H6O2 or HOCH2CH2OH Mr 62.07
CAS [107-21-1] EC No 203-473-3
Density: 1.113 g/mL at 25C (lit.) Purity > 99.5%
Measured pH Range: 5.5 - 7.8 at 25C Refractive Index Range: 1.43174 - 1.43207 at 20C
Conductivity Range: 0.1 - 0.8 mS at 25C
Synonyms: None
(C3H6O)n CAS [25322-69-4]
EC No 500-039-8 Density: 1.01 g/mL at 20C
Measured pH Range: 3.8 - 8.5 at 25C
Refractive Index Range: 1.44501 - 1.44827 at 20C Conductivity Range: 0.0 - 0.1 mS at 25C
Synonyms: 1,2,3-Propanetriol or Glycerin
HOCH2CH(OH)CH2OH or C3H8O3 Mr 92.09
CAS [56-81-5] EC No 200-289-5 Density: 1.25 g/mL (lit.)
Purity > 99.5% Measured pH Range: 6.9 - 7.4 at 25C
Refractive Index Range: 1.47220 - 1.47412 at 20C Conductivity Range: 0.0 mS at 25C
Synonyms: Polyvidone or Povidone or PVP or
Poly(1-vinyl-2-pyrrolidinone) homopolymer
(C6H9NO)n Average M
r
~10,000 CAS [9003-39-8] EC No 201-800-4
Measured pH Range: 4.2 - 5.4 at 25C
Refractive Index Range: 1.42196 - 1.42234 at 20C Conductivity Range: 476 - 812 mS at 25C
Synonyms: Diethylene oxide or Dioxane
C4H8O2 M
r
88.11
CAS [123-91-1]
EC No 204-661-8 Purity > 99.5%
Synonyms: Hexamethylene glycol
HO(CH2)6OH or C6H14O2 Mr
118.18
CAS [629-11-8] EC No 211-074-0 Purity > 97.0%
Measured pH Range: 5.3 - 8.2 at 25C
Refractive Index Range: 1.42351 - 1.42386 at 20C Conductivity Range: 0.1 - 1.5 mS at 25C
Synonyms: MPD or Hexylene glycol
CH3CH(OH)CH2C(CH3)2OH or C6H14O2
Mr
118.18 CAS [107-41-5] EC No 203-489-0
Purity > 99.0% Density: 0.925 g/mL at 25 C (lit.)
Refractive Index Range: 1.42755 - 1.42787 at 20C
Conductivity Range: 0.0 - 0.3 mS at 25C
Synonyms: sec-Propyl alcohol or Isopropanol or Isopropyl alcohol
(CH3)2CHOH or C3H8O Mr 60.10 CAS [67-63-0]
EC No 200-661-7 Purity > 99.5% Density: 0.785 g/mL at 25C (lit.)
Refractive Index Range: 1.37694 - 1.37748 at 20C
O
OH H
[ [
n
CH
3
O
OH H
3
C
[ [
n
HO
OH
OH
H
3
C CH
3
O
O
CH
3
CHCH
2
C
OH OH
CH
3
CH
3
Protein crystals with a wheat-sheaf habit.
Jennifer Wingard, University of Maryland, USA.
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S A L T S
Ammonium acetate
HR2-565 1.0 M solution 100 ml $70.00
HR2-799 8.0 M solution 200 ml $98.00
Al ......................... 0.0005%
As ...................... 0.00001%
Ba ........................ 0.0005%
Bi ......................... 0.0005%
Ca .......................... 0.001%
Cd ........................ 0.0005%
Cl ......................... 0.0005%
Co ........................ 0.0005%
Cr ......................... 0.0005%
Cu ........................ 0.0005%
Fe ........................ 0.0002%
K ............................ 0.005%
Li .......................... 0.0005%
Mg ........................ 0.0005%
Mn ........................ 0.0005%
Mo ........................ 0.0005%
Na .......................... 0.005%
Ni ......................... 0.0005%
NO3........................ 0.001%
Pb ........................ 0.0005%
SO4 ........................ 0.001%
Sr ......................... 0.0005%
Zn ........................ 0.0005%
Ammonium chloride
HR2-691 5.0 M solution 200 ml $65.00
Al ............................ 0.0005%
As ......................... 0.00001%
Ba ........................... 0.0005%
Bi ............................ 0.0005%
Ca ............................. 0.001%
Cd ........................... 0.0005%
Co ........................... 0.0005%
Cr ............................ 0.0005%
Cu ........................... 0.0002%
Fe ........................... 0.0002%
K ............................... 0.005%
Li ............................. 0.0005%
Mg ........................... 0.0005%
Mn ........................... 0.0005%
Mo ........................... 0.0005%
Na ............................. 0.005%
Ni ............................ 0.0001%
NO
3......................... 0.0005%
Pb ........................... 0.0001%
PO
4 ......................... 0.0002%
SO4 ........................... 0.002%
Sr ............................ 0.0005%
Zn ........................... 0.0002%
Ammonium phosphate monobasic
HR2-555 2.5 M solution 200 ml $68.00
Al ............................ 0.0005%
As ......................... 0.00005%
Ba ........................... 0.0005%
Bi ............................ 0.0005%
Ca ............................. 0.005%
Cd ........................... 0.0005%
Cl .............................. 0.005%
Co ........................... 0.0005%
Cr ............................ 0.0005%
Cu ........................... 0.0005%
Fe ............................. 0.002%
K ............................... 0.005%
Li ............................. 0.0005%
Mg ........................... 0.0005%
Mn ........................... 0.0005%
Mo ........................... 0.0005%
Na ............................. 0.005%
Ni ............................ 0.0005%
Pb ........................... 0.0005%
SO4 ........................... 0.005%
Sr ............................ 0.0005%
Zn ........................... 0.0005%
Ammonium fluoride
HR2-689 10.0 M solution 200 ml $71.00
Ammonium formate
HR2-659 10.0 M solution 200 ml $134.00
Al ............................ 0.0005%
As ......................... 0.00001%
Ba ........................... 0.0005%
Bi ............................ 0.0005%
Ca ............................. 0.001%
Cd ........................... 0.0005%
Cl .............................. 0.005%
Co ........................... 0.0005%
Cr ............................ 0.0005%
Cu ........................... 0.0005%
Fe ........................... 0.0005%
K ............................... 0.005%
Li ............................. 0.0005%
Mg ........................... 0.0005%
Mn ........................... 0.0005%
Mo ........................... 0.0005%
Na ............................. 0.005%
Ni ............................ 0.0005%
Pb ........................... 0.0005%
SO
4 ........................... 0.005%
Sr ............................ 0.0005%
Zn ........................... 0.0005%
Ammonium citrate dibasic
HR2-685 2.5 M solution 200 ml $95.00
Al ............................ 0.0005%
As ......................... 0.00001%
Ba ........................... 0.0005%
Bi ............................ 0.0005%
Ca ............................. 0.001%
Cd ........................... 0.0005%
Cl ............................ 0.0005%
Co ........................... 0.0005%
Cr ............................ 0.0005%
Cu ........................... 0.0005%
Fe ........................... 0.0005%
K ............................... 0.005%
Li ............................. 0.0005%
Mg ........................... 0.0005%
Mn ........................... 0.0005%
Mo ........................... 0.0005%
Na ............................. 0.005%
Ni ............................ 0.0005%
Pb ........................... 0.0005%
PO
4 ......................... 0.0005%
SO4 ........................... 0.005%
Sr ............................ 0.0005%
Zn ........................... 0.0005%
Ammonium phosphate dibasic
HR2-629 3.5 M solution 200 ml $75.00
Al ........................... 0.0005%
As ........................ 0.00001%
Ba .......................... 0.0005%
Bi ........................... 0.0005%
Ca ............................ 0.001%
Cd .......................... 0.0005%
Cl ........................... 0.0005%
Co .......................... 0.0005%
Cr ........................... 0.0005%
Cu .......................... 0.0005%
Fe .......................... 0.0005%
K .............................. 0.005%
Li ............................ 0.0005%
Mg .......................... 0.0005%
Mn .......................... 0.0005%
Mo .......................... 0.0005%
Na ............................ 0.005%
Ni ........................... 0.0005%
NO3.......................... 0.001%
Pb .......................... 0.0005%
SO
4 .......................... 0.002%
Sr ........................... 0.0005%
Zn .......................... 0.0005%
Ammonium nitrate
HR2-665 10.0 M solution 200 ml $89.00
Al ............................ 0.0005%
As ......................... 0.00001%
Ba ........................... 0.0005%
Bi ............................ 0.0005%
Ca ............................. 0.001%
Cd ........................... 0.0005%
Cl ............................ 0.0003%
Co ........................... 0.0005%
Cr ............................ 0.0005%
Cu ........................... 0.0005%
Fe ........................... 0.0002%
K ............................... 0.005%
Li ............................. 0.0005%
Mg ........................... 0.0005%
Mn ........................... 0.0005%
Mo ........................... 0.0005%
Na ............................. 0.005%
Ni ............................ 0.0005%
NO2......................... 0.0005%
Pb ........................... 0.0005%
PO
4 ......................... 0.0005%
SO4 ........................... 0.002%
Sr ............................ 0.0005%
Zn ........................... 0.0005%
Ammonium citrate tribasic pH 7.0
HR2-759 pH 7.0 - 2.5 M solution 200 ml $95.00
Synonyms: None C2H7NO2 or CH3CO2NH4
Mr 77.08 CAS [631-61-8] EC No 211-162-9 Purity > 99.0%
HR2-565: Measured pH Range: 6.8 - 7.1 at 25C
Refractive Index Range: 1.34382 - 1.34410 at 20C
Conductivity Range: 62.0 - 70.5 mS at 25C
HR2-799: Measured pH Range: 7.6 - 7.9 at 25C
Refractive Index Range: 1.40811 - 1.40885 at 20C
Conductivity Range: 68.9 - 80.0 mS at 25C
Synonyms: Formic acid ammonium salt
HCOONH4 or CH5NO2 Mr 63.06
CAS [540-69-2] EC No 208-753-9 Purity > 99.0%
Measured pH Range: 7.0 - 7.3 at 25C
Refractive Index Range: 1.40348 - 1.40398 at 20C
Conductivity Range: 579.6 - 706.0 mS at 25C
Synonyms: None NH4NO3 Mr 80.04
CAS [6484-52-2] EC No 229-347-8 Purity > 99.5%
Measured pH Range: 3.8 - 5.6 at 25C
Refractive Index Range: 1.41941 - 1.41957 at 20C
Conductivity Range: 942 - 1036 mS at 25C
Synonyms: Ammonium hydrogenphosphate or
di-Ammonium hydrogen phosphate (sec) or
Diammonium hydrogen phosphate
(NH4)2HPO4 or H9N2O4P Mr 132.06
CAS [7783-28-0] EC No 231-987-8 Purity > 99.0%
Measured pH Range: 7.8 - 8.1 at 25C Refractive Index Range: 1.40135 - 1.40362 at 20C
Conductivity Range: 87.4 - 105.7 mS at 25C
Synonyms: Ammonium dihydrogen phosphate or
Mono-ammonium phosphate or prim-Ammonium phosphate
NH4H2PO4 Mr 115.03 CAS [7722-76-1] EC No 231-764-5
Purity > 99.5% Measured pH Range: 3.5 - 3.8 at 25C
Refractive Index Range: 1.36710 - 1.36745 at 20C
Conductivity Range: 81.9 - 95.5 mS at 25C
Synonyms: Salmiac NH4Cl Mr 53.49
CAS [12125-02-9] EC No 235-186-4 Purity > 99.5%
Measured pH Range: 3.9 - 4.3 at 25C Refractive Index Range: 1.38063 - 1.38075 at 20C
Conductivity Range: 515.2 - 588.0 mS at 25C
Synonyms: Citric acid ammonium salt or
Diammonium hydrogen citrate or Di-Ammnonium citrate
C6H14N2O7 or (NH4)2C6H6O7 or HOC(CO2H)(CH2CO2NH4)2
Mr 226.19 CAS [3012-65-5] EC No 221-146-3 Purity > 99.0%
Measured pH Range: 4.7 - 4.8 at 25C
Refractive Index: 1.42003 at 20C
Conductivity Range: 76.8 - 81.9 mS at 25C
Synonyms: Citric acid triammonium salt
HOC(CO2NH4)(CH2CO2NH4)2 or C6H17N3O7
Mr 243.22 CAS [3458-72-8] EC No 222-394-5 Purity > 97.0%
HR2-759 titrated to pH 7.0 at 25C using Hydrochloric acid (HR2-581)
Refractive Index Range: 1.43010 - 1.43183 at 20C
Conductivity Range: 90.4 - 100.8 mS at 25C
Synonyms: None NH4F Mr 37.04 CAS [12125-01-8] EC No 235-185-9
Purity > 98.0% Measured pH Range: 7.4 - 8.0 at 25C Refractive Index: 1.42003 at 20C
Conductivity Range: 350.8 - 428.8 mS at 25C
O
H
3
C ONH
4
N
O O
Ag ........................... 0.0005%
Al ............................ 0.0005%
Ba ........................... 0.0005%
Bi ............................ 0.0005%
Ca ............................. 0.005%
Cd ........................... 0.0005%
Cl ............................ 0.0005%
Co ........................... 0.0005%
Cr ............................ 0.0005%
Cu ........................... 0.0005%
Fe ........................... 0.0005%
K ............................... 0.005%
Li ............................. 0.0005%
Mg ........................... 0.0005%
Mn ........................... 0.0005%
Mo ........................... 0.0005%
Na ............................. 0.005%
Ni ............................ 0.0005%
Pb ........................... 0.0005%
SiF
6............................... 0.1%
SO4 ........................... 0.005%
Sr ............................ 0.0005%
Ti ............................. 0.0005%
Zn ........................... 0.0005%
O P T I M I Z E

- R E A G E N T S
Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com 44
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Calcium chloride dihydrate
HR2-557 2.0 M solution 100 ml $74.00
As ......................... 0.00001%
Ba ........................... 0.0005%
Bi ............................ 0.0005%
Cd ........................... 0.0005%
Co ........................... 0.0005%
Cr ............................ 0.0005%
Cu ........................... 0.0005%
Fe ........................... 0.0005%
K ............................... 0.005%
Li ............................. 0.0005%
Mg ............................. 0.005%
Mn ........................... 0.0005%
Mo ........................... 0.0005%
N ............................... 0.002%
Na ............................... 0.01%
Ni ............................ 0.0005%
Pb ........................... 0.0005%
SO4 ........................... 0.005%
Sr ................................ 0.01%
Zn ........................... 0.0005%
Cesium chloride
HR2-719 1.0 M solution 100 ml $128.00
Al ............................ 0.0005%
As ......................... 0.00001%
Ba ........................... 0.0005%
Bi ............................ 0.0005%
Ca ............................. 0.005%
Cd ........................... 0.0005%
Co ........................... 0.0005%
Cr ............................ 0.0005%
Cu ........................... 0.0005%
Fe ........................... 0.0005%
K ............................... 0.005%
Li ............................. 0.0005%
Mg ........................... 0.0005%
Mn ........................... 0.0005%
Mo ........................... 0.0005%
N ............................... 0.001%
Na ............................. 0.005%
Ni ............................ 0.0005%
Pb ........................... 0.0005%
SO4 ........................... 0.002%
Sr ............................ 0.0005%
Zn ........................... 0.0005%
Cobalt(II) chloride hexahydrate
HR2-713 1.0 M solution 100 ml $64.00
Iron(III) chloride hexahydrate
HR2-717 1.0 M solution 100 ml $58.00
As ......................... 0.0005%
Ca ........................... 0.005%
Cd ........................... 0.001%
Cl ............................ 0.002%
Co ........................... 0.005%
Cr .............................. 0.01%
Cu ........................... 0.002%
Fe2
+
........................ 0.002%
K ............................. 0.005%
Mg ........................... 0.001%
Mn ............................... 0.1%
N ............................. 0.001%
Na ............................. 0.01%
Ni ............................ 0.005%
NO3........................... 0.01%
Pb ........................... 0.002%
PO4 ........................... 0.01%
SO4 ......................... 0.005%
Zn ........................... 0.002%
Hexadecyltrimethylammonium bromide
HR2-711 0.05 M solution 200 ml $61.00
Al .......................... 0.0005%
As ....................... 0.00005%
Ba ......................... 0.0005%
Bi .......................... 0.0005%
Ca ........................... 0.001%
Cd ......................... 0.0005%
Co ......................... 0.0005%
Cr .......................... 0.0005%
Cu ......................... 0.0005%
Fe ......................... 0.0005%
K ............................. 0.005%
Li ........................... 0.0005%
Mg ......................... 0.0005%
Mn ......................... 0.0005%
Mo ......................... 0.0005%
Na ........................... 0.005%
Ni .......................... 0.0005%
Pb ......................... 0.0005%
SO4 ......................... 0.005%
Sr .......................... 0.0005%
Zn ......................... 0.0005%
Cadmium sulfate hydrate
HR2-721 1.0 M solution 100 ml $88.00
As ......................... 0.00001%
Ca ............................. 0.005%
Cl .............................. 0.001%
Co ........................... 0.0005%
Cr ............................ 0.0005%
Cu ........................... 0.0005%
Fe ........................... 0.0005%
K ............................... 0.005%
Mg ........................... 0.0005%
Mn ........................... 0.0005%
N ............................. 0.0005%
Na ............................. 0.005%
Ni ............................ 0.0005%
Pb ............................. 0.002%
Zn ............................. 0.005%
Calcium acetate hydrate
HR2-567 1.0 M solution 100 ml $68.00
As ......................... 0.00001%
Ba ........................... 0.0005%
Bi ............................ 0.0005%
Cd ........................... 0.0005%
Cl .............................. 0.005%
Co ........................... 0.0005%
Cr ............................ 0.0005%
Cu ........................... 0.0005%
Fe ........................... 0.0005%
K ................................. 0.01%
Li .............................. 00005%
Mg ................................. 0.1%
Mn ........................... 0.0005%
Mo ........................... 0.0005%
Na ............................... 0.01%
Ni ............................ 0.0005%
Pb ........................... 0.0005%
SO4 ............................. 0.01%
Sr ................................ 0.02%
Zn ........................... 0.0005%
S A L T S
Ammonium sulfate
HR2-541 3.5 M solution 200 ml $68.00
Al ............................ 0.0005%
As ......................... 0.00002%
Ba ........................... 0.0005%
Bi ............................ 0.0005%
Ca ............................. 0.001%
Cd ........................... 0.0001%
Cl ............................ 0.0005%
Co ........................... 0.0005%
Cr ............................ 0.0005%
Cu ........................... 0.0002%
Fe ........................... 0.0002%
K ............................... 0.005%
Li ............................. 0.0005%
Mg ........................... 0.0005%
Mn ........................... 0.0005%
Mo ........................... 0.0005%
Na ............................. 0.005%
Ni ............................ 0.0005%
NO3........................... 0.001%
Pb ........................... 0.0002%
PO4 ......................... 0.0005%
Sr ............................ 0.0005%
Zn ........................... 0.0001%
Ammonium tartrate dibasic
HR2-679 2.0 M solution 200 ml $75.00
HR2-767 pH 7.0 - 1.6 M solution 200 ml $94.00
Al ............................ 0.0005%
As ......................... 0.00001%
Ba ........................... 0.0005%
Bi ............................ 0.0005%
Ca ............................. 0.001%
Cd ........................... 0.0005%
Cl .............................. 0.005%
Co ........................... 0.0005%
Cr ............................ 0.0005%
Cu ........................... 0.0005%
Fe ........................... 0.0005%
K ............................... 0.005%
Li ............................. 0.0005%
Mg ........................... 0.0005%
Mn ........................... 0.0005%
Mo ........................... 0.0005%
Na ............................. 0.005%
Ni ............................ 0.0005%
Pb ........................... 0.0005%
SO4 ........................... 0.005%
Sr ............................ 0.0005%
Zn ........................... 0.0005%
Cadmium chloride hydrate
HR2-715 1.0 M solution 100 ml $98.00
Synonyms: Ammonium sulphate
(NH4)2SO4 or H8N2O4S Mr 132.14
CAS [7783-20-2] EC No 231-984-1 Purity > 99.5%
Measured pH Range: 5.0 - 5.2 at 25C
Refractive Index Range: 1.39131 - 1.39166 at 20C
Conductivity Range: 313.2 - 424.2 mS at 25C
Synonyms: None CaCl2 2H2O Mr 147.01
CAS [10035-04-8] EC No 233-1408 Purity > 99.5%
Measured pH Range: 6.1 - 7.9 at 25C Refractive Index Range: 1.38108 - 1.38195 at 20C
Conductivity Range: 170.0 - 196.9 mS at 25C
Synonyms: None CsCl Mr 168.36 CAS [7647-17-8] EC No 231-600-2
Purity > 99.0% Measured pH Range: 6.0 - 6.5 at 25C
Refractive Index: 1.34591 at 20C
Conductivity Range: 106.5 - 112.9 mS at 25C
Synonyms: Cobaltous chloride hexahydrate Cl2Co2 6H2O Mr 237.93
CAS [7791-13-1] EC No 231-589-4 Purity > 98.0%
Measured pH Range: 2.1 - 3.6 at 25C
Conductivity Range: 103.4 - 114.8 mS at 25C
Synonyms: CTAB or cetrimide or cetrimonium bromide or
cetyltrimethylammonium bromide or
palmityltrimethylammonium bromide
C19H42BrN or CH3(CH2)15N(Br)(CH3)3 Mr 364.45
CAS [57-09-0] EC No 200-311-3 Purity > 99.0%
Measured pH Range: 5.3 - 5.9 at 25C
Refractive Index: 1.33573 at 20C
Conductivity Range: 1109 - 1344 mS at 25C
Synonyms: Ferric chloride hexahydrate
FeCl3 6H2O or Cl3Fe 6H2O Mr 270.30 CAS [10025-77-1] EC No 231-729-4
Measured pH Range: 0.8 - 1.0 at 25C Refractive Index: 1.37469 at 20C
Conductivity Range: 90.0 - 94.8 mS at 25C
Synonyms: L-(+)-Tartaric acid diammonium salt or
Diammonium tartrate (NH4)2C4H4O6 or C4H12N2O6 Mr 184.15
CAS [3164-29-2] EC No 221-618-9 Purity > 99.5%
HR2-679: Measured pH Range: 6.4 - 6.8 at 25C
Refractive Index Range: 1.39234 - 1.39254 at 20C
Conductivity Range: 118.2 - 134.4 mS at 25C
HR2-767: Titrated to pH 7.0 at 25C using Sodium hydroxide (HR2-583)
Refractive Index Range: 1.38139 - 1.38147 at 20C
Conductivity Range: 115.7 - 128.5 mS at 25C
Synonyms: None CdCl2 xH2O Mr 183.32 (anhyd.) CAS [654054-66-7] EC No 233-296-7
Purity 98.0% Density: 3.327 g/mL at 25C (lit.) Measured pH Range: 3.3 - 5.0 at 25C
Refractive Index Range: 1.35455 - 1.35477 at 20C Conductivity Range: 26.9 - 28.9 mS at 25C
Synonyms: Cadmium sulfate octahydrate 3CdSO4 8H2O Mr
769.52
CAS [7790-84-3] EC No 233-331-6
Purity > 99.0% (calc. based on CdSO4 8/3 H2O, KT)
Measured pH Range: 3.5 - 4.4 at 25C
Refractive Index: 1.40310 at 20C
Conductivity Range: 40.5 - 45.3 mS at 25C
Synonyms: None (CH3COO)2Ca xH2O C4H6CaO4 xH2O
Mr 158.17 (anhyd.) CAS [114460-21-8] EC No 200-540-9
Purity > 99.0% Measured pH Range: 7.6 - 7.9 at 25C
Refractive Index Range: 1.35719 - 1.35722 at 20C
Conductivity Range: 34.1 - 38.1 mS at 25C
Co Cl
Cl
O
O
-
S
O
-
O
Cd
++
45
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Lithium acetate dihydrate
Al ............................ 0.0005%
Ca ............................. 0.005%
Cl .................................. 0.5%
Cu ........................... 0.0005%
Fe ........................... 0.0005%
K ............................... 0.005%
Mg ............................. 0.001%
NH
4
+
........................... 0.05%
Na ............................. 0.005%
P ............................. 0.0005%
Pb ............................. 0.001%
SO4 ............................. 0.05%
Zn ........................... 0.0005%
HR2-669 5.0 M solution 200 ml $93.00
Lithium chloride
Al ............................ 0.0005%
As ......................... 0.00001%
Ba ............................. 0.001%
Bi ............................ 0.0005%
Ca ............................. 0.001%
Cd ........................... 0.0005%
Co ........................... 0.0005%
Cr ............................ 0.0005%
Cu ........................... 0.0005%
Fe ........................... 0.0005%
K ............................... 0.005%
Mg ........................... 0.0005%
Mn ........................... 0.0005%
Mo ........................... 0.0005%
N ............................... 0.001%
Na ............................. 0.005%
Ni .............................. 0.001%
Pb ........................... 0.0005%
SO4 ........................... 0.005%
Sr ............................ 0.0005%
Zn ........................... 0.0005%
HR2-631 10.0 M solution 200 ml $124.00
Lithium citrate tribasic tetrahydrate
Al ............................ 0.0005%
As ......................... 0.00001%
Ba ........................... 0.0005%
Bi ............................ 0.0005%
Ca ............................. 0.001%
Cd ........................... 0.0005%
Cl .............................. 0.005%
Co ........................... 0.0005%
Cr ............................ 0.0005%
Cu ........................... 0.0005%
Fe ........................... 0.0005%
K ............................... 0.005%
Mg ........................... 0.0005%
Mn ........................... 0.0005%
Mo ........................... 0.0005%
Na ............................. 0.005%
Ni ............................ 0.0005%
Pb ........................... 0.0005%
SO4 ........................... 0.005%
Sr ............................ 0.0005%
Zn ........................... 0.0005%
HR2-681 1.5 M solution 200 ml $98.00
Lithium nitrate
Al .............................. 0.001%
As ......................... 0.00001%
Ba ............................. 0.001%
Bi ............................ 0.0005%
Ca ............................... 0.01%
Cd ........................... 0.0005%
Cl .............................. 0.005%
Co ........................... 0.0005%
Cr ............................ 0.0005%
Cu ........................... 0.0005%
Fe ........................... 0.0005%
K ................................. 0.02%
Mg ........................... 0.0005%
Mn ........................... 0.0005%
Mo ........................... 0.0005%
Na ............................... 0.02%
Ni ............................ 0.0005%
Pb ........................... 0.0005%
SO
4 ............................. 0.01%
Sr ............................ 0.0005%
Zn ........................... 0.0005%
HR2-697 8.0 M solution 200 ml $85.00
HR2-545 2.0 M solution 200 ml $98.00
Al ............................ 0.0005%
As ......................... 0.00001%
Ba ........................... 0.0005%
Bi ............................ 0.0005%
Ca ............................. 0.001%
Cd ........................... 0.0005%
Cl .............................. 0.005%
Co ........................... 0.0005%
Cr ............................ 0.0005%
Cu ........................... 0.0005%
Fe ........................... 0.0005%
K ............................... 0.005%
Mg ........................... 0.0005%
Mn ........................... 0.0005%
Mo ........................... 0.0005%
Na ............................. 0.005%
Ni ............................ 0.0005%
NO3........................... 0.001%
Pb ........................... 0.0005%
Sr ............................ 0.0005%
Zn ........................... 0.0005%
S A L T S
Lithium sulfate monohydrate
Magnesium acetate tetrahydrate
HR2-561 1.0 M solution 100 ml $81.00
Al ............................ 0.0005%
As ......................... 0.00001%
Ba ........................... 0.0005%
Bi .............................. 0.001%
Ca ............................. 0.002%
Cd ........................... 0.0005%
Cl .............................. 0.005%
Co ........................... 0.0005%
Cr ............................ 0.0005%
Cu ........................... 0.0005%
Fe ........................... 0.0005%
K ............................... 0.005%
Li ............................. 0.0005%
Mn ............................. 0.002%
Mo ........................... 0.0005%
Na ............................. 0.005%
Ni ............................ 0.0005%
Pb ........................... 0.0005%
SO4 ............................. 0.02%
Sr ............................ 0.0005%
Zn ........................... 0.0005%
Synonyms: Acetic acid lithium salt CH3COOLi 2H2O
Mr 102.02 CAS [6108-17-4] EC No 208-914-3
Measured pH Range: 8.3 - 9.2 at 25C
Refractive Index: 1.38124 at 20C
Conductivity Range: 24.7 - 26.5 mS at 25C
Synonyms: Lithium chloride anhydrous
LiCl Mr 42.39 CAS [7447-41-8] EC No 231-212-3 Purity > 99.0%
Measured pH Range: 5.7 - 6.6 at 25C
Refractive Index: 1.41030 at 20C
Conductivity Range: 107.4 - 132.9 mS at 25C
Synonyms: Acetic acid magnesium salt
Mg(CH
3
COO)2Mg 4H2O or C4H6MgO4 4H2O
Mr 214.45 CAS [16674-78-5] EC No 205-554-9
Purity > 99.0% Measured pH Range: 8.1 - 8.4 at 25C
Refractive Index Range: 1.35767 - 1.35799 at 20C
Conductivity Range: 32.4 - 36.8 mS at 25C
Synonyms: None MgCl2 6H2O Mr 203.30
CAS [7791-18-6] EC No 232-094-6 Purity > 99.0%
HR2-559: Measured pH Range: 5.1 - 6.0 at 25C
Refractive Index Range: 1.37544 - 1.37603 at 20C
Conductivity Range: 136.2 - 160.2 mS at 25C
HR2-803: Measured pH Range: 3.7 - 5.7 at 25C Refractive Index: 1.43079 at 20C
Conductivity Range: 60.0 - 84.5 mS at 25C
Synonyms: Diformic acid magnesium salt C2H2O4Mg 2H2O
Mr 150.38 CAS [557-39-1] EC No 209-173-9 Purity 98.5%
Measured pH Range: 7.6 - 7.8 at 25C
Refractive Index Range: 1.35214 - 1.35241 at 20C
Conductivity Range: 45.6 - 50.3 mS at 25C
Synonyms: Magnesium nitrate
Mg(NO3)2 6H2O MgN2O6 6H2O Mr 256.41
CAS [13446-18-9] EC No 233-826-7 Purity > 99.0%
Measured pH Range: 2.7 - 3.8 at 25C
Refractive Index Range: 1.39348 - 1.39373 at 20C
Conductivity Range: 112.4 - 136.1 mS at 25C
Synonyms: Epsom salts MgSO4 7H2O Mr 246.47
CAS [10034-99-8] EC No 231-298-2 Purity > 99.5%
Measured pH: 5.6 at 25C
Conductivity Range: 51.8 mS at 25C
Synonyms: Citric acid trilithium salt or Trilithium citrate tetrahydrate
HOC(COOLi)(CH2COOLi)2 4H2O or C6H5Li3O7 4H2O
Mr 281.99 CAS [6080-58-6] EC No 213-045-8 Purity > 99.5%
Measured pH Range: 8.4 - 9.3 at 25C
Refractive Index Range: 1.38910 - 1.38970 at 20C
Conductivity Range: 30.9 - 32.5 mS at 25C
Synonyms: None LiNO3 Mr
68.95
CAS [7790-69-4] EC No 232-218-9 Purity > 99.0%
Measured pH Range: 7.0 - 7.8 at 25C
Refractive Index: 1.39391 at 20C
Conductivity Range: 139.2 - 151.5 mS at 25C
Synonyms: Lithium sulfate Li2SO4 H2O or Li2O4S H2O
Mr 127.96 CAS [10102-25-7] EC No 233-820-4 Purity > 99.0%
Measured pH Range: 3.0 - 4.3 at 25C
Refractive Index Range: 1.36520 - 1.36553 at 20C
Conductivity Range: 76.4 - 86.5 mS at 25C
Magnesium chloride hexahydrate
HR2-559 2.0 M solution 100 ml $76.00
HR2-803 5.0 M solution 200 ml $98.00
Al ............................ 0.0005%
As ......................... 0.00001%
Ba ........................... 0.0005%
Bi ............................ 0.0005%
Ca ............................. 0.005%
Cd ........................... 0.0005%
Co ........................... 0.0005%
Cr ............................ 0.0005%
Cu ........................... 0.0005%
Fe ........................... 0.0005%
K ................................... 0.2%
Li ............................. 0.0005%
Mn ........................... 0.0005%
Mo ........................... 0.0005%
N ............................. 0.0002%
Na ................................. 0.3%
Ni ............................ 0.0005%
PO4 ......................... 0.0005%
Pb ........................... 0.0005%
SO4 ........................... 0.005%
Sr ............................ 0.0005%
Zn ........................... 0.0005%
Magnesium sulfate heptahydrate
HR2-821 2.0 M solution 200 ml $102.00
Al ............................ 0.0005%
As ......................... 0.00001%
Ba ........................... 0.0005%
Bi ............................ 0.0005%
Ca ............................. 0.001%
Cd ........................... 0.0005%
Cl ............................ 0.0005%
Co ........................... 0.0005%
Cr ............................ 0.0005%
Cu ........................... 0.0005%
Fe ........................... 0.0001%
K ............................... 0.001%
Li ............................. 0.0005%
Mn ........................... 0.0005%
Mo ........................... 0.0005%
N ............................... 0.002%
Na ............................. 0.001%
Ni ............................ 0.0005%
Pb ........................... 0.0005%
Sr ............................ 0.0005%
Zn ........................... 0.0005%
Magnesium nitrate hexahydrate
HR2-657 3.0 M solution 200 ml $83.00
Al ............................ 0.0005%
As ......................... 0.00001%
Ba ............................. 0.001%
Bi .............................. 0.001%
Ca ................................. 0.5%
Cd ........................... 0.0005%
Cl .............................. 0.005%
Co ........................... 0.0005%
Cr ............................ 0.0005%
Cu ........................... 0.0005%
Fe ........................... 0.0005%
K ................................. 0.01%
Li ............................. 0.0005%
Mn ........................... 0.0005%
Mo ........................... 0.0005%
Na ............................... 0.01%
Ni ............................ 0.0005%
Pb ........................... 0.0005%
SO
4
........................... 0.005%
Sr .............................. 0.001%
Zn ........................... 0.0005%
Magnesium formate dihydrate
HR2-537 1.0 M solution 200 ml $82.00
O P T I M I Z E

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Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com 46
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S A L T S
Magnesium sulfate hydrate
HR2-633 2.5 M solution 200 ml $83.00
Ca ............................... 0.02%
Cd ............................. 0.005%
Cl ................................ 0.01%
Co ............................. 0.005%
Cu ............................. 0.005%
Fe ............................. 0.005%
K ................................. 0.05%
Na ............................... 0.01%
Ni .............................. 0.005%
Pb ............................. 0.005%
Zn ............................. 0.005%
Nickel(II) chloride hexahydrate
HR2-687 4.0 M solution 200 ml $69.00
Ca ............................. 0.005%
Cd ............................. 0.005%
Co ............................... 0.05%
Cu ............................. 0.005%
Fe ............................. 0.005%
K ................................. 0.01%
Pb ............................. 0.005%
Zn ............................. 0.005%
Potassium acetate
DNases................none detected
RNases................none detected
Proteases............none detected
Phosphatases......none detected
Pb .... 5ppm (parts per million)
HR2-671 5.0 M solution 200 ml $85.00
Potassium citrate tribasic monohydrate
HR2-683 2.5 M solution 200 ml $74.00
DL-Malic acid pH 7.0
HR2-761 pH 7.0 - 3.0 M solution 200 ml $62.00
Potassium fluoride
Al ............................ 0.0005%
As ......................... 0.00001%
Ba ........................... 0.0005%
Bi ............................ 0.0005%
Ca ............................. 0.001%
Cd ........................... 0.0005%
Cl .............................. 0.005%
Co ........................... 0.0005%
Cr ............................ 0.0005%
Cu ........................... 0.0005%
Fe ........................... 0.0005%
HF ............................. 0.001%
KOH ............................ 0.01%
K2SiF6 ...................... 0.003%
Li ............................. 0.0005%
Mg ........................... 0.0005%
Mn ........................... 0.0005%
Mo ........................... 0.0005%
Na ................................. 0.2%
Ni ............................ 0.0005%
Pb ........................... 0.0005%
SO4 ........................... 0.005%
Sr ............................ 0.0005%
Zn ........................... 0.0005%
HR2-647 6.0 M solution 100 ml $68.00
Potassium formate
Ca ............................. 0.005%
Cd ............................. 0.005%
Cl .............................. 0.005%
Co ............................. 0.005%
Cu ............................. 0.005%
Fe ............................. 0.005%
Na ................................. 0.5%
Ni .............................. 0.005%
Pb ............................. 0.005%
SO4 ........................... 0.005%
Zn ............................. 0.005%
HR2-667 14.0 M solution 200 ml $118.00
Potassium phosphate dibasic
HR2-635 4.0 M solution 200 ml $74.00
Al ............................ 0.0005%
As ......................... 0.00001%
Ba ........................... 0.0005%
Bi ............................ 0.0005%
Ca ............................. 0.001%
Cd ........................... 0.0005%
Cl .............................. 0.005%
Co ........................... 0.0005%
Cr ............................ 0.0005%
Cu ........................... 0.0005%
Fe ........................... 0.0005%
Li ............................. 0.0005%
Mg ........................... 0.0005%
Mn ........................... 0.0005%
Mo ........................... 0.0005%
Na ................................. 0.5%
Ni ............................ 0.0005%
Pb ........................... 0.0005%
SO
4 ........................... 0.005%
Sr ............................ 0.0005%
Zn ........................... 0.0005%
Synonyms: Magnesium sulphate hydrate MgSO4 xH2O
Mr 120.37 (anhyd.) CAS [22189-08-8] EC No 231-298-2 Purity > 99.0%
Measured pH Range: 6.2 - 8.2 at 25C
Refractive Index: 1.37187 at 20C
Conductivity Range: 49.2 - 59.5 mS at 25C
Synonyms: None KCl Mr 74.55
CAS [7447-40-7] EC No 231-211-8 Purity > 99.5%
Measured pH Range: 5.4 - 8.1 at 25C Refractive Index Range: 1.36868 - 1.36882 at 20C
Conductivity Range: 391.0 - 449.6 mS at 25C
Synonyms: Citric acid tripotassium salt or tri-potassium citrate
HOC(COOK)(CH2COOK)2 H2O or C6H5K3O7 H2O
Mr 324.41 CAS [6100-05-6] EC No 231-905-0 Purity > 98.0%
Measured pH Range: 8.2 - 9.9 at 25C
Refractive Index: 1.42503 at 20C
Conductivity Range: 80.6 - 91.7 mS at 25C
Synonyms: None KF or FK Mr 58.10
CAS [7789-23-3] EC No 232-151-5 Purity > 99.5%
Measured pH Range: 8.0 - 10.0 at 25C
Refractive Index: 1.35344 at 20C
Conductivity Range: 346.6 - 416.2 mS at 25C
Synonyms: Formic acid potassium salt HCOOK or CHKO2
Mr 84.12 CAS [590-29-4] EC No 209-677-9 Purity > 99.0%
Measured pH Range: 9.5 - 10.8 at 25C
Refractive Index: 1.41516 at 20C
Conductivity Range: 77.9 - 99.4 mS at 25C
Synonyms: Dipotassium hydrogenphosphate or
Dipotassium phosphate or sec.-Potassium phosphate
K2HPO4 or HK2O4P Mr 174.18 CAS [7758-11-4] EC No 231-834-5
Measured pH Range: 9.6 - 9.8 at 25C
Refractive Index Range: 1.40781 - 1.40791 at 20C
Conductivity Range: 126.9 - 148.5 mS at 25C
Synonyms: Nickel chloride NiCl2 6H2O Mr 237.69
CAS [7791-20-0] EC No 231-743-0 Purity > 98.0%
Measured pH Range: 3.1 - 3.8 at 25C
Refractive Index Range: 1.43878 - 1.43890 at 20C
Conductivity Range: 82.2 - 104.6 mS at 25C
Synonyms: ()-2-Hydroxysuccinic acid or DL-Hydroxybutanedioic acid
HO2CCH2CH(OH)CO2H or C4H6O5
Mr 134.09 CAS [6915-15-7] EC No 230-022-8 Purity > 99.0%
Titrated to pH 7.0 at 25C using Sodium hydroxide (HR2-583)
Refractive Index Range: 1.40605 - 1.40638 at 20C
Conductivity Range: 43.0 - 52.1 mS at 25C
Synonyms: None KBr or BrK Mr 119.00
CAS [7758-02-3] EC No 231-830-3 Purity > 99.5%
Measured pH Range: 8.2 - 8.6 at 25C
Refractive Index Range: 1.38454 - 1.38686 at 20C
Conductivity Range: 384.6 - 438.8 mS at 25C
Synonyms: K(acac) C2H3KO2 or CH3COOK Mr 98.14
CAS [127-08-2] EC No 204-822-2 Purity > 99.0%
Measured pH Range: 7.9 - 8.9 at 25C
Refractive Index Range: N/A
Conductivity Range: 132.3 - 146.7 mS at 25C
Potassium bromide
Al ............................ 0.0005%
As ......................... 0.00001%
Ba ........................... 0.0005%
Bi ............................ 0.0005%
BrO
3.......................... 0.001%
Ca ............................. 0.001%
Cd ........................... 0.0005%
Cl .................................. 0.1%
Co ........................... 0.0005%
Cr ............................ 0.0005%
Cu ........................... 0.0005%
Fe ........................... 0.0005%
I................................. 0.001%
Li ............................. 0.0005%
Mg ........................... 0.0005%
Mn ........................... 0.0005%
Mo ........................... 0.0005%
N ............................... 0.001%
Na ............................... 0.02%
Ni ............................ 0.0005%
Pb ........................... 0.0005%
SO4 ........................... 0.005%
Sr ............................ 0.0005%
Zn ........................... 0.0005%
HR2-779 4.0 M solution 100 ml $74.00
Potassium chloride
Al ............................ 0.0005%
As ......................... 0.00001%
Ba ............................. 0.001%
Bi ............................ 0.0005%
Br ................................ 0.05%
Ca ............................. 0.005%
Cd ........................... 0.0005%
Co ........................... 0.0005%
Cr ............................ 0.0005%
Cu ........................... 0.0005%
Fe ........................... 0.0002%
I................................. 0.002%
Li ............................. 0.0005%
Mg ............................. 0.001%
Mn ........................... 0.0005%
Mo ........................... 0.0005%
N ............................... 0.001%
Na ............................... 0.02%
Ni ............................ 0.0005%
Pb ........................... 0.0005%
PO
4 ......................... 0.0005%
SO4 ........................... 0.003%
Sr ............................ 0.0005%
Zn ........................... 0.0005%
HR2-649 4.0 M solution 200 ml $80.00
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Potassium nitrate
Al ............................ 0.0005%
As ......................... 0.00001%
Ba ........................... 0.0005%
Bi ............................ 0.0005%
Ca ............................. 0.005%
Cd ........................... 0.0005%
Cl ................................ 0.01%
Co ........................... 0.0005%
Cr ............................ 0.0005%
Cu ........................... 0.0005%
Fe ........................... 0.0002%
IO
3 .......................... 0.0005%
Li ............................. 0.0005%
Mg ............................. 0.005%
Mn ........................... 0.0005%
Mo ........................... 0.0005%
NH4
+
......................... 0.001%
Na ............................. 0.005%
Ni ............................ 0.0005%
NO2........................... 0.001%
Pb ........................... 0.0005%
PO
4 ......................... 0.0005%
SO4 ........................... 0.002%
Sr ............................ 0.0005%
Zn ........................... 0.0005%
HR2-663 3.0 M solution 200 ml $79.00
Potassium sulfate
HR2-675 0.5 M solution 200 ml $65.00
Al ............................ 0.0005%
As ......................... 0.00001%
Ba ........................... 0.0005%
Bi ............................ 0.0005%
Ca ............................. 0.005%
Cd ........................... 0.0005%
Cl ............................ 0.0005%
Co ........................... 0.0005%
Cr ............................ 0.0005%
Cu ........................... 0.0005%
Fe ........................... 0.0005%
Li ............................. 0.0005%
Mg ............................. 0.002%
Mn ........................... 0.0005%
Mo ........................... 0.0005%
N ............................. 0.0005%
Na ............................. 0.005%
Ni ............................ 0.0005%
Pb ........................... 0.0005%
PO4 ........................... 0.001%
Sr .............................. 0.005%
Zn ........................... 0.0005%
Potassium thiocyanate
HR2-695 8.0 M solution 200 ml $105.00
Al .............................. 0.001%
Ca ............................. 0.001%
Cl .............................. 0.005%
Cu ........................... 0.0005%
Fe ........................... 0.0005%
Mg ........................... 0.0005%
NH
4
+
......................... 0.003%
Na ............................. 0.005%
Pb ............................. 0.001%
SO4 ........................... 0.005%
Zn ........................... 0.0005%
Potassium sodium tartrate tetrahydrate
HR2-539 1.5 M solution 200 ml $91.00
Al ............................ 0.0005%
As ......................... 0.00001%
Ba ........................... 0.0005%
Bi ............................ 0.0005%
Ca ............................. 0.001%
Cd ........................... 0.0005%
Cl ............................ 0.0005%
Co ........................... 0.0005%
Cr ............................ 0.0005%
Cu ........................... 0.0005%
Fe ........................... 0.0005%
Li ............................. 0.0005%
Mg ........................... 0.0005%
Mn ........................... 0.0005%
Mo ........................... 0.0005%
N ............................... 0.002%
Ni ............................ 0.0005%
Pb ........................... 0.0005%
PO4 ........................... 0.001%
SO4 ........................... 0.005%
Sr ............................ 0.0005%
Zn ........................... 0.0005%
Sodium acetate trihydrate
Al ............................ 0.0005%
As ......................... 0.00001%
Bi ............................ 0.0005%
Ca ............................. 0.001%
Cd ........................... 0.0005%
Cl ............................ 0.0005%
Co ........................... 0.0005%
Cr ............................ 0.0005%
Cu ........................... 0.0005%
Fe ........................... 0.0005%
K ............................... 0.005%
Li ............................. 0.0005%
Mg ........................... 0.0005%
Mn ........................... 0.0005%
Mo ........................... 0.0005%
Ni ............................ 0.0005%
Pb ........................... 0.0005%
PO
4 ......................... 0.0005%
SO4 ........................... 0.002%
Sr ............................ 0.0005%
Zn ........................... 0.0005%
HR2-543 3.0 M solution 200 ml $72.00
HR2-763 pH 7.0 - 4.0 M solution 200 ml $65.00
Sodium bromide
HR2-699 5.0 M solution 200 ml $41.00
Ca ............................. 0.005%
Cd ............................. 0.005%
Cl .................................. 0.1%
Co ............................. 0.005%
Cu ............................. 0.005%
Fe ............................. 0.005%
K ................................... 0.2%
Ni .............................. 0.005%
Pb ............................. 0.005%
SO4 ........................... 0.005%
Zn ............................. 0.005%
L-Proline
HR2-775 1.0 M solution 100 ml $60.00
Al ............................ 0.0005%
As ......................... 0.00001%
Ba ........................... 0.0005%
Bi ............................ 0.0005%
Ca ............................. 0.001%
Cd ........................... 0.0005%
Cl ................................ 0.01%
Co ........................... 0.0005%
Cr ............................ 0.0005%
Cu ........................... 0.0005%
Fe ........................... 0.0005%
K ............................... 0.005%
Li ............................. 0.0005%
Mg ........................... 0.0005%
Mn ........................... 0.0005%
Mo ........................... 0.0005%
Na ............................. 0.005%
NH4
+
........................... .002%
Ni ............................ 0.0005%
Pb ........................... 0.0005%
SO4 ............................. 0.01%
Sr ............................ 0.0005%
Zn ........................... 0.0005%
Sodium chloride
HR2-637 5.0 M solution 200 ml $33.00
Al ............................ 0.0005%
As ......................... 0.00001%
Ba ........................... 0.0005%
Bi ............................ 0.0005%
Br .............................. 0.005%
Ca ............................. 0.001%
Cd ........................... 0.0005%
Co ........................... 0.0005%
Cr ............................ 0.0005%
Cu ........................... 0.0005%
Fe ........................... 0.0001%
I................................. 0.001%
K ............................... 0.005%
Li ............................. 0.0005%
Mg ........................... 0.0005%
Mn ........................... 0.0005%
Mo ........................... 0.0005%
N ............................... 0.001%
Ni ............................ 0.0005%
Pb ........................... 0.0005%
PO4 ......................... 0.0005%
SO4 ............................. 0.01%
Sr ............................ 0.0005%
Zn ........................... 0.0005%
Potassium phosphate monobasic
HR2-553 1.5 M solution 200 ml $69.00
Al ............................ 0.0005%
As ......................... 0.00001%
Ba ........................... 0.0005%
Bi ............................ 0.0005%
Ca ............................. 0.001%
Cd ........................... 0.0005%
Cl ............................ 0.0005%
Co ........................... 0.0005%
Cr ............................ 0.0005%
Cu ........................... 0.0005%
Fe ........................... 0.0005%
Li ............................. 0.0005%
Mg ........................... 0.0005%
Mn ........................... 0.0005%
Mo ........................... 0.0005%
N ............................... 0.001%
Na ............................. 0.005%
Ni ............................ 0.0005%
Pb ........................... 0.0005%
SO4 ........................... 0.003%
Sr ............................ 0.0005%
Zn ........................... 0.0005%
Synonyms: Monopotassium phosphate or
Potassium dihydrogen phosphate or prim.-Potassium phosphate
H2KO4P or KH2PO4 Mr 136.09 CAS [7778-77-0]
EC No 231-913-4 Purity > 99.5%
pKa
1
= 2.15 at 25C
pKa
2
= 6.82 at 25C
pKa
3
= 12.38 at 25C
Measured pH Range: 3.8 - 4.3 at 25C
Refractive Index Range: 1.35379 - 1.35414 at 20C
Conductivity Range: 67.7 - 78.1 mS at 25C
Synonyms: Potassium rhodanide KSCN or CKNS
Mr 97.18 CAS [333-20-0] EC No 206-370-1 Purity > 99.0%
Measured pH Range: 7.2 at 25C
Refractive Index Range: 1.45988 at 20C
Conductivity Range: 767.6 mS at 25C
Synonyms: (S)-pyrrolidine-2-carboxylic acid C5H9NO2
Mr 115.13 CAS [147-85-3] EC No 205-702-2 Purity > 99.5%
Measured pH Range: 6.0 - 7.0 at 25C
Refractive Index Range: 1.35260 at 20C
Conductivity Range: 5.4 - 9.1 mS at 25C
Synonyms: None KNO3 Mr 101.10
CAS [7757-79-1] EC No 231-818-8 Purity > 99.5%
Measured pH Range: 5.9 - 7.3 at 25C
Refractive Index: 1.35835 at 20C
Conductivity Range: 171.4 - 211.1 mS at 25C
Synonyms: Acetic acid sodium salt or Sodium acetate
C2H3NaO2 3H2O or CH3COONa 3H2O Mr 136.08
CAS [6131-90-4] EC No 204-823-8 Purity > 99.5%
HR2-543: Measured pH Range: 9.0 - 9.7 at 25C
Refractive Index Range: 1.36432 - 1.36440 at 20C
Conductivity Range: 73.7 - 79.9 mS at 25C
HR2-763: Titrated to pH 7.0 at 25C using Hydrochloric acid (HR2-581)
Refractive Index Range: 1.37343 - 1.37369 at 20C
Conductivity Range: 74.3 - 77.8 mS at 25C
Synonyms: None NaBr or BrNa Mr 102.89
CAS [7647-15-6] EC No 231-599-9 Purity > 99.0% Measured pH Range: 6.6 - 8.4 at 25C
Refractive Index Range: 1.39146 - 1.39846 at 20C
Conductivity Range: 309.0 - 369.4 mS at 25C
Synonyms: None NaCl Mr 58.44 CAS [7647-14-5] EC No 231-598-3 Purity > 99.5%
Measured pH Range: 5.5 - 8.6 at 25C Refractive Index Range: 1.37712 - 1.37742 at 20C
Conductivity Range: 281.2 - 354.8 mS at 25C
Synonyms: L(+)-Tartaric acid potassium sodium salt or
Rochelle salt or Seignette salt
KOCOCH(OH)CH(OH)COONa 4H2O or C
4
H
4
KNaO
6
4H2O
Mr
282.22 CAS [6381-59-5] EC No 206-156-8 Purity > 99.5%
Measured pH Range: 8.4 - 9.1 at 25C
Refractive Index Range: 1.37582 - 1.37599 at 20C
Conductivity Range: 81.1 - 97.4 mS at 25C
Synonyms: None K2SO4 Mr 174.26
CAS [7778-80-5] EC No 231-915-5 Purity > 99.0%
Measured pH Range: 6.6 - 6.8 at 25C
Refractive Index Range: N/A
Conductivity Range: 70.8 - 77.8 mS at 25C
O
N
H
OH
H
3H
2
O
CH
3
ONa C
O
HO OK P
O
O
O P T I M I Z E

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Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com 48
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S A L T S
Sodium formate
HR2-547 7.0 M solution 200 ml $68.00
HR2-765 pH 7.0 - 5.0 M solution 200 ml $69.00
Al ............................ 0.0005%
As ......................... 0.00001%
Ba ........................... 0.0005%
Bi ............................ 0.0005%
Ca ............................. 0.001%
Cd ........................... 0.0005%
Cl .............................. 0.005%
Co ........................... 0.0005%
Cr ............................ 0.0005%
Cu ........................... 0.0005%
Fe ........................... 0.0005%
K ................................. 0.01%
Li ............................. 0.0005%
Mg ........................... 0.0005%
Mn ........................... 0.0005%
Mo ........................... 0.0005%
Ni ............................ 0.0005%
Pb ........................... 0.0005%
SO4 ........................... 0.005%
Sr ............................ 0.0005%
Zn ........................... 0.0005%
Sodium fluoride
HR2-645 0.8 M solution 100 ml $63.00
Al ............................ 0.0005%
As ......................... 0.00005%
Ba ............................. 0.001%
Bi ............................ 0.0005%
Ca ............................. 0.001%
Cd ........................... 0.0005%
Cl .............................. 0.005%
Co ........................... 0.0005%
Cr ............................ 0.0005%
Cu ........................... 0.0005%
Fe ............................. 0.001%
HF ............................... 0.05%
K ................................. 0.02%
Li ............................. 0.0005%
Mg ........................... 0.0005%
Mn ........................... 0.0005%
Mo ........................... 0.0005%
NaOH .......................... 0.04%
Ni ............................ 0.0005%
Pb ........................... 0.0005%
SiF
6............................... 0.1%
SO4 ............................. 0.02%
SO3 ........................... 0.005%
Sr ............................ 0.0005%
Zn ........................... 0.0005%
Sodium malonate
HR2-747 pH 4.0 - 3.4 M solution 200 ml $79.00
HR2-749 pH 5.0 - 3.4 M solution 200 ml $79.00
HR2-751 pH 6.0 - 3.4 M solution 200 ml $79.00
HR2-707 pH 7.0 - 3.4 M solution 200 ml $79.00
HR2-807 pH 8.0 - 3.4 M solution 200 ml $79.00
HR2-809 pH 9.0 - 3.4 M solution 200 ml $79.00
Sodium nitrate
HR2-661 7.0 M solution 200 ml $75.00
Al ............................ 0.0005%
As ......................... 0.00001%
Ba ........................... 0.0005%
Bi ............................ 0.0005%
Ca ............................. 0.001%
Cd ........................... 0.0005%
Cl ............................ 0.0005%
Co ........................... 0.0005%
Cr ............................ 0.0005%
Cu ........................... 0.0005%
Fe ........................... 0.0002%
IO
3
.......................... 0.0005%
K ............................... 0.005%
Li ............................. 0.0005%
Mg ........................... 0.0005%
Mn ........................... 0.0005%
Mo ........................... 0.0005%
NH
4
+
......................... 0.002%
Ni ............................ 0.0005%
NO2........................... 0.001%
Pb ........................... 0.0005%
PO
4 ......................... 0.0002%
SO4 ........................... 0.003%
Sr ............................ 0.0005%
Zn ........................... 0.0005%
Sodium citrate tribasic dihydrate
HR2-549 1.6 M solution 200 ml $74.00
Al ............................ 0.0005%
As ......................... 0.00001%
Ba ........................... 0.0005%
Bi ............................ 0.0005%
Ca ............................. 0.005%
Cd ........................... 0.0005%
Cl .............................. 0.001%
Co ........................... 0.0005%
Cr ............................ 0.0005%
Cu ........................... 0.0005%
Fe ........................... 0.0005%
K ................................. 0.01%
Li ............................. 0.0005%
Mg ........................... 0.0005%
Mn ........................... 0.0005%
Mo ........................... 0.0005%
NH4
+
......................... 0.001%
Ni ............................ 0.0005%
Pb ........................... 0.0005%
SO4 ........................... 0.005%
Zn ........................... 0.0005%
Synonyms: Citric acid trisodium salt dihydrate or Trisodium citrate dihydrate
C6H5Na3O7 2H2O or HOC(COONa)(CH2COONa)2 2H2O
Mr 294.10 CAS [6132-04-3] EC No 200-675-3 Purity > 99.0 %
Measured pH Range: 8.2 - 8.3 at 25C
Refractive Index Range: 1.39431 - 1.39467 at 20C
Conductivity Range: 50.2 - 60.1 mS at 25C
Synonyms: Chile salpeter
NaNO3 Mr 84.99 CAS [7631-99-4] EC No 231-554-3 Purity > 99.0%
Measured pH Range: 5.0 - 6.6 at 25C
Refractive Index Range: 1.38545 - 1.38576 at 20C
Conductivity Range: 174.4 - 195.3 mS at 25C
Sodium phosphate dibasic dihydrate
HR2-639 1.0 M solution 200 ml $94.00
Al ............................ 0.0005%
As ......................... 0.00001%
Ba ........................... 0.0005%
Bi ............................ 0.0005%
Ca ............................. 0.001%
Cd ........................... 0.0005%
Cl .............................. 0.001%
Co ........................... 0.0005%
Cr ............................ 0.0005%
Cu ........................... 0.0005%
Fe ........................... 0.0005%
K ............................... 0.005%
Li ............................. 0.0005%
Mg ........................... 0.0005%
Mn ........................... 0.0005%
Mo ........................... 0.0005%
N ............................... 0.001%
Ni ............................ 0.0005%
Pb ........................... 0.0005%
SO4 ........................... 0.005%
Sr ............................ 0.0005%
Zn ........................... 0.0005%
Synonyms: sec-Sodium phosphate or di-sodium hydrogen phosphate dihydrate or
Disodium hydrogen phosphate dihydrate or Disodium phosphate
HNa2O4P 2H2O or Na2HPO4 2H2O Mr 177.99
CAS[10028-24-7] EC No 231-448-7 Purity > 99.0%
Measured pH Range: 8.7 - 8.9 at 25C
Refractive Index Range: 1.35711 - 1.35717 at 20C
Conductivity Range: 62.3 - 69.0 mS at 25C
Synonyms: None NaF Mr 41.99 CAS [7681-49-4] EC No 231-667-8 Purity > 99.0%
Measured pH Range: 8.7 - 9.9 at 25C Refractive Index Range: 1.33713 - 1.33716 at 20C
Conductivity Range: 49.9 - 52.7 mS at 25C
Synonyms: Formic acid sodium salt CHNaO2 or HCOONa
Mr 68.01 CAS [141-53-7] EC No 205-488-0 Purity > 99.0%
HR2-547: Measured pH Range: 8.4 - 9.3 at 25C
Refractive Index Range: 1.38040 - 1.38045 at 20C
Conductivity Range: 107.5 - 120.5 mS at 25C
HR2-765: Titrated to pH 7.0 at 25C using Hydrochloric acid (HR2-581)
Refractive Index Range: 1.36899 - 1.36909 at 20C
Conductivity Range: 121.8 - 134.7 mS at 25C
Synonyms: Propanedioic acid or Malonic acid
C3H4O4 (before titration with NaOH)
Mr 104.06 (before titration with NaOH) CAS [141-82-2] EC No 205-503-0 Purity > 99.0%
HR2-747: Titrated to pH 4.0 at 25C using Sodium hydroxide (HR2-583)
Refractive Index Range: 1.38388 - 1.38458 at 20C Conductivity Range: 64.2 - 74.1 mS at 25C
HR2-749: Titrated to pH 5.0 at 25C using Sodium hydroxide (HR2-583)
Refractive Index Range: 1.39285 - 1.39476 at 20C Conductivity Range: 63.5 - 77.9 mS at 25C
HR2-751: Titrated to pH 6.0 at 25C using Sodium hydroxide (HR2-583)
Refractive Index Range: 1.39940 - 1.39982 at 20C Conductivity Range: 64.0 - 75.5 mS at 25C
HR2-707: Titrated to pH 7.0 at 25C using Sodium hydroxide (HR2-583)
Refractive Index Range: 1.40091 - 1.40112 at 20C Conductivity Range: 66.8 - 77.1 mS at 25C
HR2-807: Titrated to pH 8.0 at 25C using Sodium hydroxide (HR2-583)
Refractive Index Range: 1.40121 - 1.40135 at 20C Conductivity Range: 72.6 - 73.2 mS at 25C
HR2-809: Titrated to pH 9.0 at 25C using Sodium hydroxide (HR2-583)
Refractive Index Range: 1.40119 - 1.40128 at 20C Conductivity Range: 72.2 - 73.1 mS at 25C
Sodium phosphate monobasic monohydrate
HR2-551 4.0 M solution 200 ml $94.00
As ......................... 0.00005%
Ca ............................. 0.001%
Cd ........................... 0.0005%
Cl ............................ 0.0005%
Co ........................... 0.0005%
Cr ............................ 0.0005%
Cu ........................... 0.0005%
Fe ........................... 0.0005%
K ............................... 0.005%
Mg ........................... 0.0005%
Mn ........................... 0.0005%
N ............................... 0.001%
Ni ............................ 0.0005%
Pb ........................... 0.0005%
SO4 ........................... 0.003%
Zn ........................... 0.0005%
Synonyms: Monosodium phosphate or
Sodium dihydrogen phosphate monohydrate
H2NaPO4 H2O or NaH2PO4 H2O Mr 137.99 CAS [10049-21-5]
EC No 231-449-2 Purity > 99.0%
Measured pH Range: 3.4 at 25C
Refractive Index Range: 1.38216 - 1.38229 at 20C
Conductivity Range: 58.8 - 60.5 mS at 25C
Sodium sulfate decahydrate
HR2-673 1.0 M solution 200 ml $75.00
Ca ............................. 0.002%
Cd ........................... 0.0005%
Cl ............................ 0.0005%
Co ........................... 0.0005%
Cr ............................ 0.0005%
Cu ........................... 0.0005%
Fe ........................... 0.0005%
K ............................... 0.002%
Li ............................. 0.0005%
Mg ........................... 0.0005%
Mn ........................... 0.0005%
Mo ........................... 0.0005%
N ............................. 0.0005%
Ni ............................ 0.0005%
Pb ........................... 0.0005%
PO4 ........................... 0.001%
Sr ............................ 0.0005%
Zn ........................... 0.0005%
Synonyms: Glaubers salt
Na2SO4 10H2O Mr 322.20 CAS [7727-73-3]
EC No 231-820-9 Purity > 99.0%
Measured pH Range: 6.0 - 7.8 at 25C
Refractive Index Range: 1.35184 - 1.35190 at 20C
Conductivity Range: 89.0 - 96.1 mS at 25C
O
NaO
O
O
ONa
ONa
HO
H
2
O
H
2
O
Sodium potassium phosphate (Quik Optimize)
HR2-223 Quik Optimize 100 ml, 2 bottles $175.00
Quik Optimize kit includes:
4.0 M Sodium phosphate monobasic monohydrate, 100 ml (HR2-551)
Mr 137.99 NaH2PO4 H2O CAS [10049-21-5] EC No 231-449-2
4.0 M Potassium phosphate dibasic, 100 ml (HR2-635)
Mr 174.18 K2HPO4 CAS [7758-11-4] EC No 231-834-5
Each kit contains a dilution table to create any pH (between 5.0 and 8.2) and any concentration
(between 0.2 and 4.0 M) in 0.2 increments. This kit can be used to reproduce Quik Screen
(HR2-221) reagent conditions as well as formulate optimization conditions.
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S A L T S
Succinic acid pH 7.0
HR2-709 pH 7.0 - 1.2 M solution 200 ml $94.00
Ca ............................. 0.005%
Cd ........................... 0.0005%
Cl .............................. 0.001%
Co ........................... 0.0005%
Cr ............................ 0.0005%
Cu ........................... 0.0005%
Fe ........................... 0.0005%
K ............................... 0.005%
Mg ............................. 0.001%
Mn ........................... 0.0005%
N ............................... 0.001%
Na ............................. 0.005%
Ni ............................ 0.0005%
Pb ........................... 0.0005%
PO4 ........................... 0.001%
SO4 ........................... 0.003%
Zn ........................... 0.0005%
Tacsimate pH 4, 5, 6, 7, 8, & 9
continued....Tacsimate pH 4, 5, 6, 7, 8, & 9
HR2-823 100% solution pH 4.0 200 ml $88.00
HR2-825 100% solution pH 5.0 200 ml $88.00
HR2-827 100% solution pH 6.0 200 ml $88.00
HR2-755 100% solution pH 7.0 200 ml $88.00
HR2-829 100% solution pH 8.0 200 ml $88.00
HR2-813 100% solution pH 9.0 200 ml $88.00
HR2-839 55% Tacsimate pH 6.0, 0.1 M MES pH 5.8 100 ml $95.00
HR2-843 55% Tacsimate pH 7.0, 0.1 M HEPES pH 6.8 100 ml $95.00
HR2-847 55% Tacsimate pH 8.0, 0.1 M BIS-TRIS pH 7.8 100 ml $95.00
HR2-851 55% Tacsimate pH 8.0, 0.1 M BTP pH 7.8 100 ml $95.00
Zinc acetate dihydrate
HR2-563 1.0 M solution 100 ml $74.00
Ca ............................... 0.005%
Cd ............................... 0.005%
Cl ................................ 0.005%
Co ............................... 0.005%
Cu ............................... 0.005%
Fe ............................... 0.005%
K ................................... 0.01%
Na ................................. 0.01%
Ni ................................ 0.005%
Pb ............................... 0.005%
SO4 ............................. 0.005%
Zinc sulfate heptahydrate
HR2-641 2.0 M solution 100 ml $67.00
As ......................... 0.00001%
Ca ............................. 0.001%
Cd ........................... 0.0005%
Cl ............................ 0.0005%
Co ........................... 0.0005%
Cr ............................ 0.0005%
Cu ........................... 0.0005%
Fe ........................... 0.0005%
K ............................... 0.001%
Mg ........................... 0.0005%
Mn ........................... 0.0002%
N ............................. 0.0005%
Na ............................. 0.001%
Ni ............................ 0.0005%
Pb ............................. 0.001%
Trimethylamine N-oxide dihydrate
Tryptone
HR2-777 1.0 M solution 100 ml $96.00
HR2-835 10% w/v Tryptone 100 ml $69.00
Sodium tartrate dibasic dihydrate
HR2-677 1.5 M solution 200 ml $85.00
Al ............................. 0.0005%
As .......................... 0.00001%
Ba ............................ 0.0005%
Bi ............................. 0.0005%
Ca .............................. 0.001%
Cd ............................ 0.0005%
Cl ............................. 0.0005%
Co ............................ 0.0005%
Cr ............................. 0.0005%
Cu ............................ 0.0005%
Fe ............................ 0.0005%
K ................................ 0.005%
Li .............................. 0.0005%
Mg ............................ 0.0005%
Mn ............................ 0.0005%
Mo ............................ 0.0005%
Ni ............................. 0.0005%
Pb ............................ 0.0005%
PO4 .......................... 0.0005%
SO4 ............................ 0.005%
Sr ............................. 0.0005%
Zn ............................ 0.0005%
Sodium thiocyanate
HR2-693 8.0 M solution 200 ml $75.00
Ca ................................ 0.02%
Cd .............................. 0.005%
Cl ................................. 0.05%
Co .............................. 0.005%
Cu .............................. 0.005%
Fe .............................. 0.005%
K .................................... 0.2%
Ni ............................... 0.005%
Pb .............................. 0.005%
SO
4 .............................. 0.05%
Zn .............................. 0.005%
Zinc chloride
HR2-811 2.0 M Zinc chloride 100 ml $56.00
Ca ............................. 0.001%
Cd ........................... 0.0005%
Co ........................... 0.0005%
Cr ............................ 0.0005%
Cu ........................... 0.0005%
Fe ........................... 0.0005%
K ............................... 0.001%
Mg ........................... 0.0005%
Mn ........................... 0.0005%
N ............................. 0.0005%
Na ............................. 0.001%
Ni ............................ 0.0005%
NO3............................. 0.003%
Pb ............................. 0.001%
SO4 ............................. 0.002%
Synonyms: L-(+)-Tartaric acid disodium salt or
Disodium tartrate dihydrate or Sodium tartrate dihydrate
C4H4Na2O6 2H2O Mr 230.08 CAS [6106-24-7]
EC No 212-773-3 Purity > 99.0%
Measured pH Range: 7.5 - 8.8 at 25C
Refractive Index Range: 1.37600 - 1.37625 at 20C
Conductivity Range: 67.9 - 71.0 mS at 25C
Synonyms: TMANO or N,N-Dimethylmethanamine oxide
(CH3)3NO 2H2O Mr 111.14 CAS [62637-93-8]
EC No 214-675-6 Purity > 98.0%
Measured pH Range: 8.0 - 9.3 at 25C
Refractive Index: 1.34327 at 20C
Conductivity Range: 31.0 - 67.1 mS at 25C
Synonyms: Sodium isothiocyanate or Sodium rhodanate or
Sodium rhodanide or Sodium sulfocyanate
NaSCN or CNNaS Mr 81.07 CAS [540-72-7]
EC No 208-754-4 Purity > 98.0%
Measured pH Range: 5.4 - 6.9 at 25C
Refractive Index Range: 1.45963 - 1.46046 at 20C
Conductivity Range: 137.1 - 161.5 mS at 25C
Synonyms: Zinc diacetate
Zn(CH3COO)2 2H2O or C4H6O4Zn 2H2O Mr 219.52
CAS [5970-45-6] EC No 209-170-2 Purity > 98.0%
Measured pH Range: 5.5 - 5.8 at 25C
Refractive Index Range: 1.35739 - 1.35749 at 20C
Conductivity Range: 16.0 - 17.6 mS at 25C
Synonyms: Zinc chloride anhydrous ZnCl
2
Mr
136.30
CAS [7646-85-7] EC No 231-592-0 Purity > 98.0%
Measured pH Range: 3.1 - 4.1 at 25C
Refractive Index Range: 1.37596 - 1.37597 at 20C
Conductivity Range: 90.2 - 104.1 mS at 25C
Synonyms: None ZnSO4 7H2O Mr 287.56
CAS [7446-20-0] EC No 231-793-3 Purity > 99.5%
Measured pH Range: 3.4 - 4.0 at 25C
Refractive Index Range: 1.38259 - 1.38264 at 20C
Conductivity Range: 54.1 - 57.4 mS at 25C
Synonyms: Dicarboxylic acid C4. or Butanedioic acid
C4H6O4 or HOOCCH2CH2COOH Mr 118.09 CAS [110-15-6]
EC No 203-740-4 Purity > 99.5%
pKa
1
= 4.2 at 25C pKa
2
= 5.6 at 25C
Titrated to pH 7.0 at 25C using Sodium hydroxide (HR2-583)
Refractive Index Range: 1.36373 - 1.36407 at 20C
Conductivity Range: 70.1 - 79.7 mS at 25C
O
NaO
O
ONa
OH
2H
2
O
OH H
H
2H
2
O Zn
2+
[ [
2
O
O H
3
C
-
S
N
Na
O
O
S
O
O
Zn
++
Tacsimate is a pH titrated mixture of organic acids: 1.8305 M Malonic acid,
0.25 M Ammonium citrate tribasic, 0.12 M Succinic acid, 0.3 M DL-Malic acid,
0.4 M Sodium acetate trihydrate, 0.5 M Sodium formate, 0.16 M Ammonium tartrate dibasic
HR2-823: Titrated to pH 4.0 at 25C using Sodium hydroxide (HR2-583)
Refractive Index Range: 1.38590 - 1.38655 at 20C Conductivity Range: 73.1 - 79.8 mS at 25C
HR2-825: Titrated to pH 5.0 at 25C using Sodium hydroxide (HR2-583)
Refractive Index Range: 1.39432 - 1.39476 at 20C Conductivity Range: 76.2 - 83.1 mS at 25C
HR2-827: Titrated to pH 6.0 at 25C using Sodium hydroxide (HR2-583)
Refractive Index Range: 1.39981 - 1.40017 at 20C Conductivity Range: 77.4 - 79.9 mS at 25C
HR2-755: Titrated to pH 7.0 at 25C using Sodium hydroxide (HR2-583)
Refractive Index Range: 1.40102 - 1.40171 at 20C Conductivity Range: 75.7 - 90.7 mS at 25C
Continued next column....
HR2-829: Titrated to pH 8.0 at 25C using Sodium hydroxide (HR2-583)
Refractive Index Range: 1.40129 - 1.40166 at 20C Conductivity Range: 74.9 - 79.8 mS at 25C
HR2-813: Titrated to pH 9.0 at 25C using Sodium hydroxide (HR2-583)
Refractive Index Range: 1.40142 - 1.40151 at 20C Conductivity Range: 75.6 - 75.9 mS at 25C
HR2-839: Titrated to pH 5.8 at 25C using Hydrochloric acid (HR2-581)
Refractive Index Range: 1.37622 at 20C Conductivity Range: 94.2 mS at 25C
HR2-843: Titrated to pH 6.8 at 25C using Hydrochloric acid (HR2-581)
Refractive Index Range: 1.37822 at 20C Conductivity Range: 92.6 - 96.0 mS at 25C
HR2-847: Titrated to pH 7.8 at 25C using Hydrochloric acid (HR2-581)
Refractive Index Range: 1.37808 at 20C Conductivity Range: 93.8 mS at 25C
HR2-851: Titrated to pH 7.8 at 25C using Hydrochloric acid (HR2-581)
Refractive Index Range: 1.37949 at 20C Conductivity Range: 93.7 mS at 25C
Measured pH Range: 7.0 - 7.1 at 25C
Refractive Index: 1.35052 at 20C
Conductivity Range: 7.7 - 8.5 mS at 25C
O P T I M I Z E

- R E A G E N T S
Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com 50
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BIS-TRIS propane pH 7.0 Buffer
HR2-795 pH 7.0 - 1.0 M solution 100 ml $119.00
HEPES sodium Buffer (Useful pH Range: 6.6 - 8.5)
HR2-577 1.0 M solution 100 ml $98.00
HR2-733 pH 7.5 - 1.0 M solution 100 ml $98.00
HR2-931-01 pH 6.8 - 1.0 M solution 185 ml $206.00
HEPES Buffer (Useful pH Range: 6.8 - 8.2)
HR2-585 1.0 M solution 100 ml $84.00
HR2-785 pH 7.0 - 1.0 M solution 100 ml $84.00
HR2-729 pH 7.5 - 1.0 M solution 100 ml $84.00
Citric acid pH 3.5 Buffer (Useful pH Range: 2.2 - 6.5)
Citric acid BIS-TRIS propane Buffer (Useful pH Range: 2.5 - 9.5)
HR2-757 pH 3.5 - 1.0 M solution 100 ml $32.00
HR2-831 1.0 M Citric acid 100 ml $33.00
HR2-833 1.0 M BIS-TRIS propane 100 ml $119.00
Al ............................ 0.0005%
As ......................... 0.00001%
Ba ........................... 0.0005%
Bi ............................ 0.0005%
Ca ............................. 0.001%
Cd ........................... 0.0005%
Cl ............................ 0.0005%
Co ........................... 0.0005%
Cr ............................ 0.0005%
Cu ........................... 0.0005%
Fe ........................... 0.0005%
K ............................... 0.005%
Li ............................. 0.0005%
Mg ........................... 0.0005%
Mn ........................... 0.0005%
Mo ........................... 0.0005%
Na ............................. 0.005%
Ni ............................ 0.0005%
Oxalate (C2O4) ........... 0.05%
Pb ........................... 0.0005%
PO4 ......................... 0.0005%
SO4 ........................... 0.002%
Sr ............................ 0.0005%
Tartrate (C4H4O6) ......... 0.2%
Zn............................. 0.0005%
B U F F E R S
ADA Buffer (Useful pH Range: 5.6 - 7.5)
HR2-507 0.5 M solution 100 ml $88.00
HR2-817 pH 6.5 - 1.0 M solution 100 ml $98.00
Al ............................ 0.0005%
As ......................... 0.00001%
Ba ........................... 0.0005%
Bi ............................ 0.0005%
Ca ............................. 0.001%
Cd ........................... 0.0005%
Cl .................................. 0.1%
Co ........................... 0.0005%
Cr ............................ 0.0005%
Cu ........................... 0.0005%
Fe ........................... 0.0005%
K ............................... 0.005%
Mg ........................... 0.0005%
Mn ........................... 0.0005%
Mo ............................. 0.005%
Na ............................... 0.05%
Ni ............................ 0.0005%
Pb ........................... 0.0005%
SO4 ........................... 0.005%
Sr ............................ 0.0005%
Zn ........................... 0.0005%
BICINE Buffer (Useful pH Range: 7.4 - 9.3)
HR2-509 1.0 M solution 100 ml $76.00
HR2-723 pH 9.0 - 1.0 M solution 100 ml $76.00
BIS-TRIS Buffer
Ca............................... 0.001%
Cd ........................... 0.0005%
Cl .............................. 0.005%
Co ........................... 0.0005%
Cr ............................ 0.0005%
Cu ........................... 0.0005%
Fe ........................... 0.0005%
K ............................... 0.005%
Mg ........................... 0.0005%
Mn ........................... 0.0005%
Na ............................. 0.005%
Ni ............................ 0.0005%
Pb ........................... 0.0005%
SO4 ........................... 0.005%
Zn ........................... 0.0005%
Synonyms: N-(2-Acetamido)iminodiacetic acid or
N-(Carbamoylmethyl)iminodiacetic acid
C6H10N2O5 or H2NCOCH2N(CH2CO2H)2 Mr 190.16
CAS [26239-55-4] EC Number 247-530-0 Purity > 99.0%
pKa = 6.6 at 25C
HR2-507: Titrate to useful pH range using Sodium hydroxide (HR2-583)
Refractive Index: 1.35012 at 20C
Conductivity Range: 23.9 - 28.6 mS at 25C
HR2-817: Titrated to pH 6.5 at 25C using Sodium hydroxide (HR2-583)
Refractive Index Range: 1.36776 - 1.36803 at 20C
Conductivity Range: 42.7 - 43.8 mS at 25C
Synonyms: Citric acid anhydrous
C6H8O7 or HOC(COOH)(CH2COOH)2
Mr 192.12 CAS [77-92-9] EC No 201-069-1 Purity > 99.5%
pKa1 = 3.13 at 25C pKa2 = 4.76 at 25C pKa3 = 6.4 at 25C
HR2-757: Titrated to pH 3.5 at 25C using Sodium hydroxide (HR2-583)
Refractive Index Range: N/A
Conductivity Range: 28.7 - 33.3 mS at 25C
Citrate BIS-TRIS propane (CBTP) is a buffer system useful across pH 2.5 to 9.5. Using CBTP
one can use a single buffer system to screen pH 2.5 to 9.5 by varying the ratio of Citric acid to
BIS-TRIS propane. Although the two buffer reagents are made available separately for conve-
nient, individual replacement, they are designed to be used together, with the supplied CBTP
buffer titration table to create a crystallization buffer system covering pH 2.5 to 9.5.
HR2-831: Measured pH Range: 1.2 - 1.5 at 25C
Refractive Index: 1.35682 at 20C Conductivity Range: 7.8 - 8.7 mS at 25C
HR2-833: Measured pH Range: 11.2 - 11.3 at 25C
Refractive Index: 1.38076 at 20C Conductivity Range: 309.0 - 364.0 mS at 25C
Synonyms: 4-(2-hydroxyethyl)piperazine-1-ethanesulfonic acid or
N-(2-hydroxyethyl)piperazine-N'-(2-ethanesulfonic acid) or
2-[4-(2-Hydroxyethyl)-1-piperazinyl]ethanesulfonic acid
C8H18N2O4S Mr 238.31 CAS [7365-45-9] EC No 230-907-9
Purity > 99.0% pKa = 7.5 at 25C
HR2-585: Titrate to useful pH range using Sodium hydroxide (HR2-583)
Measured pH Range: 5.5 - 5.5 at 25C
Refractive Index Range: 1.37121 - 1.37186 at 20C
Conductivity Range: 161.2 - 178.4 mS at 25C
HR2-785: Titrated to pH 7.0 at 25C using Sodium hydroxide (HR2-583)
Refractive Index Range: 1.37161 at 20C Conductivity Range: 7.1 mS at 25C
HR2-729: Titrated to pH 7.5 at 25C using Sodium hydroxide (HR2-583)
Refractive Index Range: 1.37184 at 20C Conductivity Range: 12.1 mS at 25C
Synonyms: HEPES sodium salt or
4-(2-hydroxyethyl)piperazine-1-ethanesulfonic acid sodium salt or
N-(2-hydroxyethyl)piperazine-N'-(2-ethanesulfonic acid) sodium salt
C8H17N2NaO4S Mr 260.29 CAS [75277-39-3] EC No 278-169-7
Purity > 99.5% pKa = 7.5 at 25C
HR2-577: Titrate to useful pH range using Hydrochloric acid (HR2-581)
Measured pH Range: 10.5 - 10.9 at 25C
Refractive Index Range: N/A
Conductivity Range: 25.9 - 28.2 mS at 25C
HR2-733: Titrated to pH 7.5 at 25C using Hydrochloric acid (HR2-581)
Refractive Index Range: 1.37713 - 1.37743 at 20C
Conductivity Range: 40.6 - 42.1 mS at 25C
HR2-931-01: HEPES sodium pH 6.8 - 1.0 M solution is a Custom Shop reagent and is used to
reproduce and optimize Crystallization Reagents for use with the Silver Bullets kits.
Synonyms: N,N-Bis(2-hydroxyethyl)glycine
C6H13NO4 Mr 163.17 CAS [150-25-4] EC No 205-755-1
Purity > 99.0% pKa = 8.3 at 25C
HR2-509: Titrate to useful pH range using Sodium hydroxide (HR2-583)
Measured pH Range: 5.1 - 5.3 at 25C
Refractive Index Range: 1.35805 - 1.35821 at 20C
Conductivity Range: 114.3 - 114.7 mS at 25C
HR2-723: Titrated to pH 9.0 at 25C using Sodium hydroxide (HR2-583)
Refractive Index Range: 1.36078 - 1.36104 at 20C
Conductivity Range: 24.3 - 24.7 mS at 25C
Synonyms: 2,2-Bis(hydroxymethyl)-2,2',2"-nitrilotriethanol or
2-Bis(2-hydroxyethyl)amino-2-(hydroxymethyl)-1,3-propanediol or
Bis(2-hydroxyethyl)amino-tris(hydroxymethyl)methane
C8H19NO5 Mr 209.24 CAS [6976-37-0] EC No 230-237-7
Purity > 99.0% pKa = 6.4 at 20C pKa = 6.5 at 25C
HR2-781: Titrated to pH 5.5 at 25C using Hydrochloric acid (HR2-581)
Refractive Index Range: 1.37130 - 1.37150 at 20C
Conductivity Range: 38.4 - 43.1 mS at 25C
HR2-783: Titrated to pH 6.5 at 25C using Hydrochloric acid (HR2-581)
Refractive Index Range: 1.36860 - 1.36926 at 20C
Conductivity Range: 22.9 - 28.7 mS at 25C
HR2-906-24: BIS-TRIS pH 7.8 - 1.0 M solution is a Custom Shop reagent and is used to
reproduce and optimize Crystallization Reagents for use with the Silver Bullets kits.
Synonyms: 1,3-Bis[tris(hydroxymethyl)methylamino]propane
CH2[CH2NHC(CH2OH)3]2 or C11H26N2O6
Mr 282.34 CAS [64431-96-5] EC No 264-899-3
Purity > 99.0% pKa1 = 6.8 at 25C pKa2 = 9.0 at 25C
HR2-795: Titrated to pH 7.0 at 25C using Hydrochloric acid (HR2-581)
Refractive Index Range: 1.39139 - 1.39143 at 20C
Conductivity Range: 41.7 - 42.5 mS at 25C
N
OH
O
HO
O
NH2 O
N N
S OH OH
O
O
HR2-781 pH 5.5 - 1.0 M solution 100 ml $124.00
HR2-783 pH 6.5 - 1.0 M solution 100 ml $124.00
HR2-906-24 pH 7.8 - 1.0 M solution 185 ml $151.00
51
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B U F F E R S
MES monohydrate Buffer (Useful pH Range: 5.2 - 7.1)
HR2-587 0.5 M solution 100 ml $98.00
HR2-787 pH 6.5 - 1.0 M solution 100 ml $98.00
HR2-943-07 pH 5.8 - 1.0 M solution 185 ml $138.00
Ca ............................. 0.005%
Cd ........................... 0.0005%
Cl .............................. 0.005%
Co ........................... 0.0005%
Cr ............................ 0.0005%
Cu ........................... 0.0005%
Fe ........................... 0.0005%
K ............................... 0.005%
Mg ........................... 0.0005%
Mn ........................... 0.0005%
Na ............................. 0.005%
Ni ............................ 0.0005%
Pb ........................... 0.0005%
SO4 ........................... 0.005%
Zn ........................... 0.0005%
Synonyms: MES or 2-(N-morpholino)ethanesulfonic acid or
4-morpholineethanesulfonic acid monohydrate
C6H13NO4S H2O Mr 213.25 CAS [145224-94-8]
EC No 224-632-3 Purity > 99.0% pKa = 6.1 at 25C
HR2-587: Titrate to useful pH range using Sodium hydroxide (HR2-583)
Measured pH Range: 3.2 - 3.4 at 25C
Refractive Index Range: 1.34690 - 1.34731 at 20C
Conductivity Range: 166.4 - 259.0 mS at 25C
HR2-787: Titrated to pH 6.5 at 25C using Sodium hydroxide (HR2-583)
Refractive Index Range: 1.36197 - 1.36229 at 20C
Conductivity Range: 19.2 - 25.6 mS at 25C
HR2-943-07: MES monohydrate pH 5.8 - 1.0 M solution is a Custom Shop reagent and is used to
reproduce and optimize Crystallization Reagents for use with the Silver Bullets kits.
Sodium cacodylate trihydrate Buffer (Useful pH Range: 5.0 - 7.4)
HR2-575 1.0 M solution 100 ml $178.00
HR2-737 pH 6.5 - 1.0 M solution 100 ml $178.00
Synonyms: Cacodylic acid sodium salt trihydrate or
Dimethylarsinic acid sodium salt or Dimethylarsonic acid sodium salt
C2H6AsNaO2 3H2O or (CH3)2AsO2Na 3H2O Mr 214.03
CAS [6131-99-3] EC No 204-708-2 pKa = 6.2 at 25C Purity > 98.0%
HR2-575: Titrate to useful pH range using Hydrochloric acid (HR2-581)
Measured pH Range: 8.3 - 8.5 at 25C
Refractive Index: 1.35596 at 20C
Conductivity Range: 33.3 - 37.7 mS at 25C
HR2-737: Titrated to pH 6.5 at 25C using Hydrochloric acid (HR2-581)
Refractive Index Range: 1.35651 - 1.35707 at 20C
Conductivity Range: 43.5 - 45.4 mS at 25C
Imidazole Buffer (Useful pH Range: 6.2 - 7.8)
HR2-573 1.0 M solution 100 ml $68.00
HR2-819 pH 7.0 - 1.0 M solution 100 ml $82.00
Ca ............................. 0.001%
Cd ........................... 0.0005%
Cl .............................. 0.005%
Co ........................... 0.0005%
Cr ............................ 0.0005%
Cu ........................... 0.0005%
Fe ........................... 0.0005%
K ............................... 0.005%
Mg ........................... 0.0005%
Mn ........................... 0.0005%
Na ............................. 0.005%
Ni ............................ 0.0005%
Pb ........................... 0.0005%
SO4 ................................ 0.005%
Zn ........................... 0.0005%
Synonyms: 1,3-Diaza-2,4-cyclopentadiene or Glyoxaline
C3H4N2 Mr 68.08 CAS [288-32-4] EC No 206-019-2
Purity > 99.5% pKa = 6.95 at 25C
HR2-573: Titrate to useful pH range using Hydrochloric acid (HR2-581)
Measured pH Range: 10.2 - 10.6 at 25C
Refractive Index: 1.34423 at 20C
Conductivity Range: 72.2 - 85.2 mS at 25C
HR2-819: Titrated to pH 7.0 at 25C using Hydrochloric acid (HR2-581)
Refractive Index Range: 1.34847 - 1.34872 at 20C
Conductivity Range: 46.1 - 49.2 mS at 25C
Sodium citrate tribasic dihydrate Buffer (Useful pH Range: 3.0 - 6.2)
HR2-571 1.0 M solution 100 ml $41.00
HR2-735 pH 5.6 - 1.0 M solution 100 ml $41.00
Al ............................ 0.0005%
As ......................... 0.00001%
Ba ........................... 0.0005%
Bi ............................ 0.0005%
Ca ............................. 0.005%
Cd ........................... 0.0005%
Cl .............................. 0.001%
Co ........................... 0.0005%
Cr ............................ 0.0005%
Cu ........................... 0.0005%
Fe ........................... 0.0005%
K ................................. 0.01%
Li ............................. 0.0005%
Mg ........................... 0.0005%
Mn ........................... 0.0005%
Mo ........................... 0.0005%
NH
4
+
......................... 0.001%
Ni ............................ 0.0005%
Pb ........................... 0.0005%
SO
4 ................................ 0.005%
Zn ........................... 0.0005%
Synonyms: Citric acid trisodium salt dihydrate or
Trisodium citrate dihydrate C6H5Na3O7 2H2O or
HOC(COONa)(CH2COONa)2 2H2O Mr 294.10
CAS [6132-04-3] EC No 200-675-3 Purity > 99.0%
pKa1 = 3.1 at 25C pKa2 = 4.8 at 25C pKa3 = 5.4 at 25C
HR2-571: Titrate to useful pH range using Hydrochloric acid (HR2-581)
Measured pH Range: 8.1 - 8.3 at 25C Refractive Index: 1.37370 at 20C
Conductivity Range: 60.0 - 67.5 mS at 25C
HR2-735: Titrated to pH 5.6 at 25C using Hydrochloric acid (HR2-581)
Refractive Index Range: 1.37423 - 1.37502 at 20C
Conductivity Range: 67.4 - 72.4 mS at 25C
O
NaO
O
O
ONa
ONa
HO
H
2
O
H
2
O
Sodium acetate trihydrate Buffer (Useful pH Range: 3.6 - 5.6)
HR2-569 1.0 M solution 100 ml $29.00
HR2-789 pH 4.5 - 1.0 M solution 100 ml $29.00
HR2-731 pH 4.6 - 1.0 M solution 100 ml $29.00
Al ............................ 0.0005%
As ......................... 0.00001%
Bi ............................ 0.0005%
Ca ............................. 0.001%
Cd ........................... 0.0005%
Cl ............................ 0.0005%
Co ........................... 0.0005%
Cr ............................ 0.0005%
Cu ........................... 0.0005%
Fe ........................... 0.0005%
K ............................... 0.005%
Li ............................. 0.0005%
Mg ........................... 0.0005%
Mn ........................... 0.0005%
Mo ........................... 0.0005%
Ni ............................ 0.0005%
Pb ........................... 0.0005%
PO4 ......................... 0.0005%
SO4 ........................... 0.002%
Sr ............................ 0.0005%
Zn ........................... 0.0005%
Synonyms: Acetic acid sodium salt C2H3NaO2 3H2O or
CH3COONa2 3H2O M
r
136.08 CAS [6131-90-4]
EC No 204-823-8 Purity > 99.5% pKa = 4.8 at 25C
HR2-569: Titrate to useful pH range using Hydrochloric acid (HR2-581)
Measured pH Range: 8.7 - 9.2 at 25C
Refractive Index Range: 1.34405 - 1.34408 at 20C
Conductivity Range: 48.0 - 51.7 mS at 25C
HR2-789: Titrated to pH 4.5 at 25C using Hydrochloric acid (HR2-581)
Refractive Index Range: 1.34558 - 1.34564 at 20C
Conductivity Range: 60.8 - 67.7 mS at 25C
HR2-731: Titrated to pH 4.6 at 25C using Hydrochloric acid (HR2-581)
Refractive Index Range: 1.34553 - 1.34556 at 20C
Conductivity Range: 59.3 - 63.4 mS at 25C
3H
2
O
CH
3
ONa C
O
Tris Buffer (Useful pH Range: 7.0 - 9.0)
HR2-589 1.0 M solution 100 ml $44.00
HR2-725 pH 8.5 - 1.0 M solution 100 ml $44.00
Al ............................ 0.0005%
As ......................... 0.00001%
Ba ........................... 0.0005%
Bi ............................ 0.0005%
Ca ............................. 0.001%
Cd ........................... 0.0005%
Cl ............................ 0.0005%
Co ........................... 0.0005%
Cr ............................ 0.0005%
Cu ........................... 0.0005%
Fe ........................... 0.0005%
K ............................... 0.005%
Li ............................. 0.0005%
Mg ........................... 0.0005%
Mn ........................... 0.0005%
Mo ........................... 0.0005%
Na ............................. 0.005%
Ni ............................ 0.0005%
Pb ........................... 0.0005%
SO
4
......................... 0.0005%
Sr ............................ 0.0005%
Zn ........................... 0.0005%
Synonyms: Trizma

base or THAM or Tris base or

2-Amino-2-(hydroxymethyl)-1,3-propanediol or
Tris(hydroxymethyl)aminomethane or Trometamol
NH2C(CH2OH)3 or C4H11NO3 Mr 121.14 CAS [77-86-1]
EC No 201-064-4 Purity > 99.8% pKa = 8.1 at 25C
HR2-589: Titrate to useful pH range using Hydrochloric acid (HR2-581)
Measured pH Range: 10.9 - 11.0 at 25C
Refractive Index Range: 1.35044 - 1.35048 at 20C
Conductivity Range: 197.0 - 223.0 mS at 25C
HR2-725: Titrated to pH 8.5 at 25C using Hydrochloric acid (HR2-581)
Refractive Index Range: 1.35364 - 1.35369 at 20C
Conductivity Range: 18.1 - 21.5 mS at 25C
O P T I M I Z E

- R E A G E N T S
Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com 52
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Synonyms: TRIS HCl or Trizma

hydrochloride or
Tris(hydroxymethyl)aminomethane hydrochloride
NH2C(CH2OH)3 HCl or C4H11NO3 HCl
Mr 157.60 CAS [1185-53-1] EC No 214-684-5
Purity > 99.0% pKa = 8.1 at 25C
HR2-579: Titrate to useful pH range using Sodium hydroxide (HR2-583)
Measured pH Range: 3.9 - 4.1 at 25C Refractive Index: 1.35882 at 20C
Conductivity Range: 49.1 - 53.3 mS at 25C
HR2-727: Titrated to pH 8.5 at 25C using Sodium hydroxide (HR2-583)
Refractive Index: 1.35961 at 20C Conductivity: 60.3 mS at 25C
TRIS hydrochloride Buffer (Useful pH Range: 7.0 - 9.0)
HR2-579 1.0 M solution 100 ml $45.00
HR2-727 pH 8.5 - 1.0 M solution 100 ml $45.00
Al ............................ 0.0005%
As ........................... 0.0001%
Ba ........................... 0.0005%
Bi ............................ 0.0005%
Ca ............................. 0.001%
Cd ........................... 0.0005%
Co ........................... 0.0005%
Cr ............................ 0.0005%
Cu ........................... 0.0005%
Fe ........................... 0.0005%
K ............................... 0.005%
Li ............................. 0.0005%
Mg ........................... 0.0005%
Mn ........................... 0.0005%
Mo ........................... 0.0005%
Na ............................. 0.005%
Ni ............................ 0.0005%
Pb ........................... 0.0005%
SO
4
........................... 0.005%
Sr ............................ 0.0005%
Zn ........................... 0.0005%
B U F F E R S
Hydrochloric acid
HR2-581 1.0 M solution 100 ml $27.00
Ag ....................... 0.000001%
Al ........................ 0.000005%
As ..................... 0.0000005%
Ba ....................... 0.000002%
Bi .......................... 0.00001%
Br .............................. 0.005%
Ca ......................... 0.00005%
Cd ....................... 0.000001%
Co ....................... 0.000001%
Cr ........................ 0.000002%
Cu ....................... 0.000002%
Fe ......................... 0.00002%
Hg ....................... 0.000001%
K ........................... 0.00001%
Li ......................... 0.000001%
Mg ......................... 0.00001%
Mn ....................... 0.000001%
Mo ....................... 0.000002%
Na ......................... 0.00005%
NH4
+
....................... 0.0001%
Ni ........................ 0.000002%
PO
4 ......................... 0.0005%
Pb ....................... 0.000002%
SO4 ......................... 0.0001%
SO3 ......................... 0.0001%
Sr ........................ 0.000001%
TI ........................ 0.000005%
Zn ....................... 0.000005%
Synonyms: None HCl Mr
36.46 CAS [7647-01-0] EC No 231-595-7
Conductivity Range: 320.4 - 378.2 mS at 25C
Sodium hydroxide
HR2-583 1.0 M solution 100 ml $27.00
Al ............................ 0.0005%
Ca ........................... 0.0005%
Cd ........................... 0.0005%
Cl .............................. 0.005%
Cu ........................... 0.0005%
Fe ........................... 0.0005%
K ................................. 0.02%
Mg ........................... 0.0005%
P ............................. 0.0005%
Pb ............................. 0.001%
SO
4 ............................. 0.05%
Zn ........................... 0.0005%
Synonyms: Caustic soda NaOH or HNaO Mr 40.00 CAS [1310-73-2]
EC No 215-185-5 Purity > 98.0%
d e s c r i p t i o n
Non Detergent Sulfobetaines (NDSB) are a new family of non denaturing
protein solubilizing agents with a wide range of applications. NDSBs have
been used for protein extraction, solubilization, and crystallization. NDSBs
are zwitterionic, they can be removed by dialysis since they do not form
micelles, and they do not alter the biophysical properties of biological
buffers.
This particular class of reagent was developed by Laurent Vuillard
(University of Cambridge, Department of Pathology, UK) in collaboration
with T. Rabilloud (CENG, Grenoble, France).
References:
1. Vuillard, L., T. Rabilloud, M.E. Goldberg, A New Additive for Protein Crystallization., FEBS Letters., 1994.,
353(3):, 294-296.
2. Vuillard, L., et al., Protein crystallography with non detergent sulfobetaines., J. Cryst. Growth. (1996) 168,
150-154.
3. Vuillard, L., Rabilloud, T., Goldberg, M.E., Eur. J. Biochem., (1998) 256, 1, 128-135.
4. http://www.path.cam.ac.uk/~lv213/
Crystal picture of RPA in an earring motif.
Courtesy of Gloria Borgstahl.
University of Toledo
Department of Chemistry
NDSB-195
HR2-703 powder 5 g $98.00
NDSB-201
HR2-701 powder 5 g $27.00
NDSB-221
HR2-791 powder 5 g $133.00
NDSB-211
HR2-793 powder 5 g $181.00
NDSB-256
HR2-705 powder 5 g $98.00
S O L U B I L I Z I N G A G E N T S
w H a T a r e n d s b s ?
Synonyms: Dimethylethylammonium propane sulfonate or
Non-detergent sulfobetaine C
7
H
17
NO
3
S Mr 195.3
Purity > 95.0% by TLC
Synonyms: 3-(1-Pyridino)-1-propane sulfonate
C
8
H
11
NO
3
S Mr 201.2
CAS Number 15471-17-7 EC No 2394913
Purity > 99.0%
Synonyms: Dimethyl-2(-Hydroxyethyl)-(3-Sulfopropyl)-ammonium,
Inner Salt or Non detergent sulfobetaine 211
C
7
H
17
NO
4
S Mr 211.28 Purity > 99.0%
Synonyms: Non-detergent sulfobetaine 221 or
3-(1-Methylpiperidinium)-1-propane Sulfonate
C
9
H
19
NO
3
S Mr 221.3 Purity > 99.0%
Synonyms: Dimethylbenzylammonium propane sulfonate
C
12
H
19
NO
3
S Mr 257.4
Purity > 99.0% by TLC
53
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d e s c r i p t i o n
Try using TCEP hydrochloride instead of dithiothreitol (DTT) as a
reducing agent in your crystallization setups. TCEP hydrochloride
is soluble in water to 310 g per L. TCEP hydrochloride is Tris
(2-carboxyethyl)phosphine hydrochloride. It is an odorless (non-
volatile) reducing agent that is more stable and effective than DTT
or 2-Mercaptoethanol. Unlike DTT, TCEP hydrochloride retains
its reducing power at acid pHs (pH 5) and at pHs above 7.5. It
is unreactive toward other functional groups found in proteins.
Unlike DTT (dithiothreitol), TCEP hydrochloride does not contain a free thiol, and therefore does
not require removal before reaction with a thiol-reactive reagents. It reduces disulfides but apparently
not mercurythiol bonds. TCEP hydrochloride is compatible with many heavy atoms and may be used
during heavy atom derivatization. It is more stable at a higher pH and at higher temperatures than is
DTT and for a longer period of time in buffers without metal chelators such as EGTA.
References
1. Tris(2-carboxyethyl)phosphine stabilization of RNA: comparison with dithiothreitol for use with nucleic acid and thiophosphoryl chemistry. Rhee SS, Burke DH.
Anal Biochem 325, 137-43 (2004) PN51541.
2. A comparison between the sulfhydryl reductants tris(2-carboxyethyl)phosphine and dithiothreitol for use in protein biochemistry. Getz EB, Xiao M, Chakrabarty
T, Cooke R, Selvin PR. Anal Biochem 273, 73-80 (1999) PN34481
3. Burns, J.A., et al. (1991). Selective reduction of disulfides by tris-(2-carboxyethyl)-phosphine. J. Org. Chem. 56, 2648-2650
4. Oda , Y., et al. (2001). Nature Biotech19, 379-382
n
Stable reducing agent for crystallization trials

n
Highly soluble
n
Odorless and non-volatile
n
Effective at acidic and alkaline pH

a p p l i c a t i o n
f e a t u r e s
Order Information
Cat. No. Name Description Price
HR2-651 TCEP hydrochloride 1 g $46.00
HR2-801 TCEP hydrochloride 10 g $290.00
P OH
O
HO
O
OH O
HCl

H
2
O
2 RSH
R R S S
P OH
O
HO
O
OH O
HCl

O
T C e P H y d r o C H l o r I d e
s I l I C a H y d r o G e l

O P T I M I Z E

- R E D U C I N G A G E N T
Hanging drop vapor diffusion crystal.
Ertugrul Cansizoglu, University of Texas, Southwestern, USA.
O P T I M I Z E

- C R Y O P R O T E C T A N T S
Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com 54
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Paratone-N is a viscous cryoprotectant for both small and large
molecule crystallography.
1
Mildly air-unstable, small molecule com-
pounds can be coated with Paratone-N under an inert atmosphere.
The Paratone-N protected crystal sample can be cryocooled in a
chilled nitrogen gas stream. Paratone-N has also been used suc-
cessfully as a cryoprotectant for biological macromolecule crystals.
It is used during cryocrystallography to displace and reduce the
amount of water (mother liquor, reagent) on the crystal after the
crystal is mounted in a cryoloop. Coating the crystal with Paratone-N can minimize evaporation from
the crystal and reduce exposure and slow diffusion of air (oxygen) to the crystal.
Paratone-N is also known as Parabar 10312, Paratone 8277, and Infineum V8512.
References
1. H. Hope, Cryocrystallography of biological macromolecules: a generally applicable method. Acta Cryst. (1988) B44, 22-26.
2. Structure of the ligand-binding domain (LBD) of human androgen receptor in complex with a selective modulator LGD2226. F. Wang, X.- Liu, H. Li, K.- Liang,
J. N. Miner, M. Hong, E. A. Kallel, A. van Oeveren, L. Zhi and T. Jiang. Acta Cryst. (2006). F62, 1067-1071
3. H. Hope, Annu. Rev. Biophys. Chem. 1990 19:107-126
4. S. Parkin and H. Hope, J. Appl. Cryst. (1998) pages 945-953
n
Cryoprotectant

a p p l i c a t i o n
Order Information
Cat. No. Name Description Price
HR2-643 Paratone-N 100 ml $55.00
P A R A T O N E - N
d e s c r i p t i o n
Used during cryocrystallography to displace and reduce the
amount of water (mother liquor, reagent) on the crystal after the
crystal is mounted in a cryoloop. Coating the crystal with oil can
minimize evaporation from the crystal and reduce exposure and
slow diffusion of air (oxygen) to the crystal.
References
1. Structure of the open form of Aspergillus nidulans 3-dehydroquinate synthase at 1.7 resolution. C.E.
Nichols, A.R. Hawkins and D.K. Stammersa. Acta Crystallographic Section D, Volume 60, Part 5, Pages 971-973, May 2004.
2. H. Hope, Annu. Rev. Biophys. Chem. 1990 19:107-126
n
Cryoprotectant

n
Perfluoropolyether oil
n
Low viscosity
n
Low surface tension
a p p l i c a t i o n
f e a t u r e s
Order Information
Cat. No. Name Description Price
HR2-814 Perfluoropolyether PFO-X175/08 1 ml $25.00
P e r F l u o r o P o l y e T H e r P F o - x 1 7 5 / 0 8
Trypsin jungle.
Allan DArcy, Switzerland.
s I l I C a H y d r o G e l

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- O I L S
d e s c r i p t i o n
Floating Drop Vapor Diffusion
The floating drop vapor diffusion technique
1
has been described
using Fluorinert FC-70. The transparent and high density FC-70
liquid is not miscible with most crystallization reagents, samples,
oil, or water. When Fluorinert liquid and the sample/reagent are
mixed, the sample/reagent drop immediately separates from
the Fluorinert and floats on top of the Fluorinert (see figure 1).
Crystals grown using the floating drop technique do not stick
to the crystallization plate. As the sample/reagent/crystal is not
miscible with the Fluorinert liquid, complete separation of the crystal from the Fluorinert is straight
forward. Crystals grown using the floating drop technique can be removed from the drop without
mechanical damage. Drop volumes can vary widely using the floating drop method. Handling of the
crystal is improved.
Floating and Stirring Technique (FAST)
The floating and stirring technique (FAST)
2
has been described using Fluorinert FC-70 liquid. The
method, which involves a sample and reagent mixture applied over a non-miscible dense liquid
(Fluorinert FC-70) without contact and slow stirring can also be combined with slow cooling or warm-
ing (See figure 2). The floating and stirring method has been reported to accelerate growth of the
crystal as well as prevent subsequent spontaneous nucleation. The method has been used with seed
crystals to promote the growth of a larger crystal without the appearance of subsequent crystals.
Fluorinert: FC-70 Fluid
Mr: 820
CAS number: [86508-42-1]
Pour point: -25C
Boiling point: 215C
Nonflammable
Vapor pressure: 15 pascals
Liquid density: 1,940 kg/m3
Kinematic viscosity: 12 centistokes
Absolute viscosity: 24 centipoise
Coeffecient of expansion: 0.0010C-1
Surface tension: 18 dynes/cm
Refractive index: 1.303
Water solubility: 8 ppmw
Ozone depletion potential: 0
Dielectric strength: 40 kV, 0.1 gap
Dielectric constant: 1.98
Electrical resistivity: 2.3 x 1015 ohm cm
Appearance: Clear, colorless
References
1. Application of a two-liquid system to sitting-drop vapour-diffusion protein crystallization. Adachi, H. et al, Acta Cryst. (2003) D59, 194-196
2. Promotion of large protein crystal growth with stirring solution. Adachi, H. et al. Jpn. J. Appl. Phys. Vol. 41 (2002) pp.1025-1027
3. Two-liquid hanging-drop vapour-diffusion technique of protein crystallization. Hiroaki Adachi et al. Japanese Journal of Applied Physics.
Vol. 43, No. 1A/B, 2004, pp.L79-L81.
n
Drop floating crystallization

n
Viscosity similar to water with approximately
75% greater density
n
Stable
n
Completely fluorinated
a p p l i c a t i o n
f e a t u r e s
Order Information
Cat. No. Name Description Price
HR2-797 100% Fluorinert FC-70 Fluid 100 ml $99.00
F L U O R I N E R T

F C - 7 0 F L U I D
figure 2
Floating and stirring technique
Fluorinert
Protein Solution
Stir Bar
figure 1
Floating drop vapor diffusion technique
Crystallization Drop
Reservoir Solution
Fluorinert
O P T I M I Z E

- O I L S
Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com 56
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d e s c r i p t i o n
Oils used for microbatch and modified microbatch crystallization
under oil.
Al's Oil is a 50:50 (volume:volume) mixture of Paraffin Oil and
Silicon Oil.
Paraffin oil
Synonym: Mineral oil
CAS Number [8042-47-5]
EC Number 2324558
RTECS PY8047000
Appearance Colorless, clear, viscous liquid
Refractive Index n20/D 1.467 (literature) n20/D 1.468
IR Spectroscopy Grade
Density 0.85 g/ml at 20C
Viscosity 40-42 CST at 25C
References
1. The advantages of using a modified microbatch method for rapid screening of protein crystallization conditions. Allan D'Arcy et al. Acta Cryst. (2003).
D59, 396-399
Containerless crystallization is a micromethod of batch crystalliza-
tion of proteins under conditions where the sample and precipi-
tant have no contact with the surface of the crystallization plate.
Drops (2 to 20 l) of sample combined with crystallization reagent
are pipetted at the interface between two layers of inert and immis-
cible silicon oils contained in a 24, 48, or 96 well plate and sealed.
Plates and plain cover slides are available separately. It is easier to
position drops into the center of the oil using plates with larger
reservoirs (i.e. 24 and 48 well plates) but this requires more oil. When using the 96 well plates, drops of
higher density (polymer based reagents such as PEG) tend to migrate to the side walls of the plate.
The method relies upon the use of two immiscible oils (FMS and DMS). Poly-3, 3, 3-Trifluoropropyl-
methylsiloxane (FMS), a branched silicone compound, has a higher density than Polydimethylsiloxane
(DMS). Therefore, one can create a bilayer with a distinct interface by placing the two oils in a reservoir.
Neither DMS nor FMS is miscible with water. The oils are compatible with most crystallization reagents
including but not limited to buffers, salts, polymers, and MPD. Additional protein or crystallization
reagents can be added to the drop in the oil. Crystal mounting as well as seeding can be performed
while the drop is in the oil. During crystal mounting, the drop will maintain a spherical shape and
decrease in size as sample is withdrawn. During seeding, crystal seeds will sediment to the bottom of
the drop. It has been observed that large, perfect, single crystals can be grown using the containerless
crystallization method. Crystals typically nucleate at the oil/water interface and grow into the drop.
n
Microbatch crystallization
n
Screen different temperatures without
condensation
n
Protect the sample from oxidation
n
Under oil crystallization

n
Contact free crystallization by microbatch
under oil
n
Utilizes two immiscible oils to create a
simulated containerless system
a p p l i c a t i o n
a p p l i c a t i o n
f e a t u r e s
f e a t u r e s
Order Information
Cat. No. Name Description Price
HR3-411 Paraffin Oil 100% - 250 ml $40.00
HR3-421 Paraffin Oil 100% - 1 L $100.00
HR3-415 Silicon Oil 100% - 250 ml $150.00
HR3-423 Silicon Oil 100% - 1 L $350.00
HR3-413 Al's Oil (50:50 Paraffin:Silicon) - 250 ml $90.00
HR3-417 Combo Oil Pack (Paraffin, Als, & Silicon Oil) - 250 ml of each $225.00
Order Information
Cat. No. Name Description Price
HR2-593 DMS Oil 100 ml $73.00
HR2-595 FMS Oil 100 ml $171.00
M I C r o b a T C H C r y s T a l l I z a T I o n o I l s
FMS Oil (Lower):
M
r
: 2350
D: 1.25
cSt: 300
Ref Index: 1.381
DMS Oil (Upper):
M
r
: 410
D: 0.873
cSt: 2
Ref Index: 1.390
C O N T A I N E R L E S S C R Y S T A L L I Z A T I O N
CH
3
CH
3
CH
3
F
CH
3 Si Si Si
Si
CH
3
CH
3
CH
3
CH
3
O O CH
3
F F
( (
n
CH
3
CH
3
CH
3
CH
3 Si Si Si
CH
3
CH
3
CH
3
O O CH
3
( (
n
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Gels are a very efficient media for growing macromolecular crys-
tals.
1-5
Silica gels in particular have the advantage in that they are
stable, usable over a wide range of temperatures (0-60C), and are
compatible with a wide variety of precipitants and additives used
for crystal growth.
Gels can reduce nucleation and sedimentation, provide added sta-
bility, and allow crystals to grow larger. The porous network mini-
mizes natural convection. Crystals are suspended in the gel net-
work so that they do not form sediments and can grow free from
strain exerted by the container or other crystals. Heterogeneous
and secondary nucleation are reduced in the presence of a Silica Hydrogel.
3
The Silica Hydrogel can
be used for liquid-gel, liquid-gel-liquid, vapor diffusion, as well as dialysis crystallization methodolo-
gies. The gel is compatible with a wide range of salts, polymers, organic solvents, and buffers used for
macromolecular crystallization in a pH range from 3 to 10.
Each Silica Hydrogel kit contains 12 tubes of Sodium silicate solution and 12 tubes of Acetic acid solu-
tion, 500 l each. All solutions are sterile filtered and formulated using ultra-pure water. Crystallization
accessories are sold separately.
References
1. Robert, M.C. & Lefaucheux, F., J. Crystal Growth (1988) 90, 358.
2. Provost, K. & Robert, M.C., J. Crystal Growth (1991) 110, 258.
3. M.C. Robert, K. Provost, & F. Lefaucheux, Crystallization of Nucleic Acids and Protein, A Practical Approach, Oxford Univ Press (1992) 127-143.
4. McPherson A., Methods in Enzymology (1985) 114, 112.
5. Cudney, B., Patel, S., McPherson, A., Acta Cryst. (1994) D50, 479-483.
n
Quick & easy kit format for crystallization in
gels
n
Proprietary Silica Hydrogel formulation with
neutral pH
n
Gel matrix can reduce nucleation &
sedimentation
a p p l i c a t i o n
f e a t u r e s
Order Information
Cat. No. Name Description Price
HR2-310 Silica Hydrogel 500 l, 24 tubes $95.00
O P T I M I Z E

- s I l I C a H y d r o G e l k I T
d e s c r i p t i o n
Low melting (LM) agaroses are the result of a derivatization process
by organic synthesis. The main properties of these agaroses are
their low melting and gelling temperatures when compared with
standard agaroses. LM agaroses have higher clarity (gel transpar-
ency) than gels of standard agaroses. The gelling temperature of
LM agaroses is 24 to 28C.
The structure of the polysaccharide is that of a galactan, formed by linking agarobioses by links 1-3, 1-4.
This chemical structure gives agaroses the capacity to form strong gels even at low temperatures. The
gels have a macroreticular structure with a very open mesh which can be adjusted simply by varying
the concentration of the agarose. The macroreticule structure of the agarose gel is formed by hydrogen
bonds, which makes the gel reversible, transforming the gel into a solution by heating. The absence
of ionic groups makes the gel a neutral structure. With no interaction, macromolecules can migrate
through the gel mesh, making the gel an efficient sieve for biological macromolecules.
n
Crystallization in agarose gel
n
Gel matrix can reduce nucleation and sedi-
mentation
n
Crystallization grade
n
Low melting agarose
a p p l i c a t i o n
f e a t u r e s
Order Information
Cat. No. Name Description Price
HR8-092 LM Agarose 10 g $58.00
O P T I M I Z E

- L M A G A R O S E
O P T I M I Z E

- I Z I T C R Y S T A L D Y E

Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com 58
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Differentiate protein crystals from salt crystals
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0.22 micron sterile filtered solution
n
Small molecule dye penetrates solvent
channels of macromolecular crystals,
coloring the crystals blue.
n
Salt crystals cannot absorb Izit and remain
colorless

a p p l i c a t i o n
f e a t u r e s
Order Information
Cat. No. Name Description Price
HR4-710 Izit Crystal Dye 0.5 ml vial $15.00
A crystal! Is it protein or is it salt? Have you ever asked yourself this
question? Sure you could mount the crystal in question and take a
quick look at the diffraction pattern but perhaps youre too lazy or
you dont have an x-ray facility. Well, theres always the crush test.
One simply takes a MicroTool or Crystal Probe and crushes
the suspect crystal. A click or solid crunch is indicative of salt while
a powder or silent destruction might indicate one just destroyed a
perfectly good protein crystal. Izit is here to help. Simply place one
l of Izit in the sample drop and wait for an hour or so. Izit is a small
molecule dye which will fill the solvent channels in protein crystals,
coloring the crystals blue. With the appropriate dilution, Izit will in
fact leave a clear drop with blue crystals, as illustrated in the picture
to the right. Salt crystals do not possess these large solvent channels.
Therefore, Izit cannot enter the crystal, leaving one with a clear crystal
and a blue drop. Izit is especially nice for small microcrystals or ques-
tionable precipitate. The 0.5 ml vial of Izit is sufficient for more than
thousands of crystallization drops.
Each Izit vial contains 0.5 ml of Izit dye. All solutions are formulated using ultra-pure water and are sterile
filtered. Crystallization accessories are sold separately.
d e s c r i p t i o n
Picturesque crystal of multidrug efflux pump AcrB.

Markus Seeger, University Zrich, Switzerland.
Nuclear receptor crystal.
Gilbert Bey, ALIX, Illkirch, France.
Crystals of Thermus thermophilus enolase.
From the group of Paola Fucini,
Max Planck Institute for Molecular Genetics, Berlin, Germany.
Protein crystals.
Kasumi Kobayashi, Taiji Nakae and Hiroyuki Akama.
The Kitasato Institute, Kanagawa, Japan.
s t o c k o p t i o n s

k i t s
Protein crystal.
Alexey Rak, Max-Planck-Institut fysiologie, Department of Physical Biochemistry, Dortmund, Germany.
T A B L E O F C O N T E N T S
P A G E S
62 s t o c k o p t i o n s

s a l t
63 s t o c k o p t i o n s

p h
64 - 67 s t o c k o p t i o n s

b u f f e r
68 s t o c k o p t i o n s

- c r y o p r o

n
Crystallization grade salt reagent stock
solutions for screen formulation and
optimization
n
Preformulated and sterile filtered
n
Easy transition from screening to optimization
n
Synergistic with Hampton Research screens,
kits, & reagents
n
49 unique salts, including organic acids
n
Highly concentrated, ready to dilute
a p p l i c a t i o n
f e a t u r e s
d e s c r i p t i o n
StockOptions reagent kits are reagent tool boxes for the
macromolecular crystal grower. They offer precisely for-
mulated, high quality crystallization grade reagent stocks
in convenient, cost-effective kits. The chemicals utilized
in these kits are the same crystallization grade, ultra-high
purity chemicals utilized in the Hampton Research kits such
as Crystal Screen and Crystal Screen 2. StockOptions
reagents are carefully formulated under strict quality stan-
dards to ensure reliable performance and lot-to-lot consis-
tency.
Each reagent in a StockOptions kit is available in convenient concentration, making crystal setups quick
and easy. Gone is the tedious task of finding and sourcing reagents, as well as costly and time-consuming
reagent formulation.
Preformulated reagents also reduce the activation energy between the discovery of preliminary screen
conditions and the task of setting experiments for optimization. The generation of custom screens or
optimized conditions now simply involves pipetting StockOptions reagents from convenient kits.
StockOptions Salt contains 49 unique salts, preformulated at convenient stock concentrations, each in 10
ml volumes, all in a single kit with a 5" x 9" footprint that saves precious lab space. It is designed to help
researchers improve the speed, accuracy, precision, and quality of the formulation of crystallization opti-
mization solutions. Researchers can use the individual StockOptions reagents to formulate custom screen
solutions or accurately reproduce standard screen solutions from Hampton Research crystallization kits.
All one needs to do is select the reagent and pipet.
The convenience also reduces the chance of errors. Preformulated stocks remove calculation, measure-
ment, and formulation errors. No more second guessing how the reagent was formulated, what specific
chemical was used, when it was made, or how to precisely reproduce that reagent when it is gone and
more reagent is required for additional setups.
StockOptions are cost-effective, time saving reagents. When a StockOptions kit is purchased, one is
using reagents as preformulated stocks in reasonable volumes. You buy only the reagents you need, not
a large container of raw material that may sit out on shelves for years to come. Waste is further reduced
since there is no chance for formulation or measuring errors. When you need larger volumes, Hampton
Research offers individual, preformulated, sterile filtered Optimize crystallization reagents which include
salts, polymers, organics solvents, and buffers. Each of the 49 salts offered in StockOptions Salt is available
individually as an Optimize crystallization grade reagent. Optimize, Custom Shop, StockOptions, and all
Hampton Research kits are synergistic research tools.
StockOptions kits also lower the costs associated with making crystallization reagents since there is no
need to purchase sterile filters, filtration devices, or pre-sterilized storage containers. Cost savings are also
extended to labor since time can now be better utilized for sample production and purification or setting
crystallization experiments.
References
1. Purification, crystallization and preliminary crystallographic characterization of the caspase-recruitment domain of human Nod1. T. Srimathi, S. L. Robbins,
R. L. Dubas, J.-H. Seo and Y. C. Park. Acta Cryst. (2007). F63, 21-23.
s T o C k o P T I o n s

S A L T
Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com 62
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Order Information
Each StockOptions Salt kit contains 49 unique reagents.
Cat. No. Name Description Price
HR2-245 StockOptions Salt 10 ml, tube format $285.00
stockoptions salt kit formulation
1.0 M Ammonium acetate
5.0 M Ammonium chloride
2.5 M Ammonium phosphate monobasic
10.0 M Ammonium fluoride
10.0 M Ammonium formate
2.5 M Ammonium citrate dibasic
3.5 M Ammonium phosphate dibasic
10.0 M Ammonium nitrate
3.5 M Ammonium sulfate
2.0 M Ammonium tartrate dibasic
1.0 M Calcium acetate hydrate
2.0 M Calcium chloride dihydrate
5.0 M Lithium acetate dihydrate
10.0 M Lithium chloride
1.5 M Lithium citrate tribasic tetrahydrate
8.0 M Lithium nitrate
2.0 M Lithium sulfate monohydrate
1.0 M Magnesium acetate tetrahydrate
2.0 M Magnesium chloride hexahydrate
1.0 M Magnesium formate dihydrate
3.0 M Magnesium nitrate hexahydrate
2.5 M Magnesium sulfate hydrate
4.0 M Nickel(II) chloride hexahydrate
5.0 M Potassium acetate
4.0 M Potassium chloride
2.5 M Potassium citrate tribasic monohydrate
1.5 M Potassium phosphate monobasic
6.0 M Potassium fluoride
14.0 M Potassium formate
3.0 M Potassium phosphate dibasic
3.0 M Potassium nitrate
1.5 M Potassium sodium tartrate tetrahydrate
0.5 M Potassium sulfate
8.0 M Potassium thiocyanate
3.0 M Sodium acetate trihydrate
5.0 M Sodium chloride
1.6 M Sodium citrate tribasic dihydrate
5.0 M Sodium phosphate monobasic monohydrate
0.8 M Sodium fluoride
7.0 M Sodium formate
1.0 M Sodium phosphate dibasic dihydrate
3.4 M Sodium malonate pH 7.0
7.0 M Sodium nitrate
1.0 M Sodium sulfate decahydrate
1.5 M Sodium tartrate dibasic dihydrate
8.0 M Sodium thiocyanate
1.2 M Succinic acid pH 7.0
1.0 M Zinc acetate dihydrate
2.0 M Zinc sulfate heptahydrate
Reagent
Number
1
2
3
4
5
6
7
8
9
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
32
33
34
35
36
37
38
39
40
41
42
43
44
45
46
47
48
49
Reagent
n
Crystallization grade buffer stocks for crystal
screening, optimization, & production
n
Titrated, preformulated, & ready-to-use buffers
n
pH 2.2 - 11.0
n
Easy transition from screening to optimization
and production
n
Synergistic with Hampton Research screens,
kits, & reagents
n
Concentrated stocks
n
Sterile filtered
a p p l i c a t i o n
f e a t u r e s
d e s c r i p t i o n
StockOptions reagent kits are reagent tool boxes for the
macromolecular crystal grower. They offer precisely for-
mulated, high quality crystallization grade reagent stocks
in convenient, cost-effective kits. The chemicals utilized
in these kits are the same crystallization grade, ultra-high
purity chemicals utilized in the Hampton Research kits such
as Crystal Screen and Crystal Screen 2. StockOptions
reagents are carefully formulated under strict quality stan-
dards to ensure reliable performance and lot-to-lot con-
sistency.
StockOptions pH is a convenient and complete kit with buffers pHed from 2.2 to 11.0 in increments of
0.2 pH units. Each reagent is available in convenient concentration, making crystal setups quick and easy.
Gone is the tedious task of finding and sourcing reagents, as well as costly and time-consuming reagent
formulation.
The generation of custom screens or optimized conditions now simply involves pipetting StockOptions
pH reagents from one convenient kit. This portfolio of buffers between pH 2.2 and 11, along with wide
arrays of salts, polymers, and organic solvents also stimulates creativity since all of the tools are readily and
conveniently available. All one needs to do is select the reagent and pipet.
The convenience also reduces the chance of errors. Preformulated stocks remove calculation, measure-
ment, and formulation errors. No more second guessing how the reagent was formulated, what specific
chemical was used, when it was made, or how to precisely reproduce that reagent when it is gone and
more reagent is required for additional setups.
StockOptions pH is cost-effective. When a StockOptions kit is purchased, one is using reagents as prefor-
mulated stocks in reasonable volumes. You buy only the reagents you need, not a large container of raw
material that may sit out on shelves for years to come. When you need larger volumes, Hampton Research
offers individual, preformulated, sterile filtered Optimize crystallization reagents which include salts,
polymers, organics solvents, and buffers. Optimize, Custom Shop, StockOptions, and all Hampton
Research kits are synergistic research tools.
References
1. Structure of the ribosomal interacting GTPase YjeQ from the enterobacterial species Salmonella typhimurium. D. K. Stammers et al. Acta Cryst. (2007).
F63, 922928
2. Increasing the size of microcrystals by fine sampling of pH limits. A. McPherson. J. Appl. Cryst. (1995). 28, 362-365.
63
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Order Information
Each StockOptions pH kit contains 45 unique reagents. To order individual reagents, use Custom
Shop

catalog number listed below. Refer to page 37 for further details.

Cat. No. Name Description Price
HR2-241 StockOptions pH 10 ml, tube format $275.00
HR2-941-** StockOptions pH Custom Shop 185 ml
Reagents 1-39 $139.00
Reagents 40-45 $151.00
** = reagent number 1-45
s T o C k o P T I o n s

P
H
sToCkoPTIons PH kIT ForMulaTIon
pH
2.2
2.4
2.6
2.8
3.0
3.2
3.4
3.6
3.8
4.0
4.2
4.4
4.6
4.8
5.0
5.2
5.4
5.6
5.8
6.0
6.2
6.4
6.6
6.8
7.0
7.2
7.4
7.6
7.8
8.0
8.2
8.4
8.6
8.8
9.0
9.2
9.4
9.6
9.8
10.0
10.2
10.4
10.6
10.8
11.0
Reagent
1.0 M Citric acid
1.0 M Citric acid
1.0 M Citric acid
1.0 M Citric acid
1.0 M Citric acid
1.0 M Citric acid
1.0 M Citric acid
1.0 M Citric acid
1.0 M Citric acid
1.0 M Sodium acetate trihydrate
1.0 M Sodium acetate trihydrate
1.0 M Sodium acetate trihydrate
1.0 M Sodium acetate trihydrate
1.0 M Sodium acetate trihydrate
1.0 M Sodium citrate tribasic dihydrate
1.0 M Sodium citrate tribasic dihydrate
1.0 M Sodium citrate tribasic dihydrate
1.0 M Sodium citrate tribasic dihydrate
1.0 M Sodium citrate tribasic dihydrate
1.0 M Sodium cacodylate trihydrate
1.0 M Sodium cacodylate trihydrate
1.0 M Sodium cacodylate trihydrate
1.0 M Sodium cacodylate trihydrate
1.0 M Sodium cacodylate trihydrate
1.0 M HEPES sodium
1.0 M HEPES sodium
1.0 M HEPES sodium
1.0 M HEPES sodium
1.0 M HEPES sodium
1.0 M TRIS hydrochloride
1.0 M TRIS hydrochloride
1.0 M TRIS hydrochloride
1.0 M TRIS hydrochloride
1.0 M TRIS hydrochloride
0.5 M CAPSO
0.5 M CAPSO
0.5 M CAPSO
0.5 M CAPSO
0.5 M CAPSO
0.5 M CAPS
0.5 M CAPS
0.5 M CAPS
0.5 M CAPS
0.5 M CAPS
0.5 M CAPS
Titrant
Sodium hydroxide
Sodium hydroxide
Sodium hydroxide
Sodium hydroxide
Sodium hydroxide
Sodium hydroxide
Sodium hydroxide
Sodium hydroxide
Sodium hydroxide
Hydrochloric acid
Hydrochloric acid
Hydrochloric acid
Hydrochloric acid
Hydrochloric acid
Hydrochloric acid
Hydrochloric acid
Hydrochloric acid
Hydrochloric acid
Hydrochloric acid
Hydrochloric acid
Hydrochloric acid
Hydrochloric acid
Hydrochloric acid
Hydrochloric acid
Hydrochloric acid
Hydrochloric acid
Hydrochloric acid
Hydrochloric acid
Hydrochloric acid
Sodium hydroxide
Sodium hydroxide
Sodium hydroxide
Sodium hydroxide
Sodium hydroxide
Sodium hydroxide
Sodium hydroxide
Sodium hydroxide
Sodium hydroxide
Sodium hydroxide
Sodium hydroxide
Sodium hydroxide
Sodium hydroxide
Sodium hydroxide
Sodium hydroxide
Sodium hydroxide
Reagent
Number
1
2
3
4
5
6
7
8
9
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
32
33
34
35
36
37
38
39
40
41
42
43
44
45
n
Focused, titrated buffer stocks for crystal
screening, optimization & production
n
pH range 2.2 - 6.5 (44 unique reagents)
n
0.1 pH unit increments for fine tuning pH
n
Compatible with Hampton Research screens,
kits & reagents
n
1.0 M concentrated stocks
Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com 64
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StockOptions Sodium Acetate buffer kit is a preformulated, sterile filtered set of titrated buffer stocks.
The reagents are supplied as 1.0 M stock solutions in 10 ml volumes. Each StockOptions Sodium
Acetate buffer reagent is carefully titrated using Hydrochloric acid. The kit is comprised of 21 unique
reagents covering the pH range of 3.6 to 5.6 in 0.1 pH unit increments.
References
1. Crystal structures of reduced, oxidized, and mutated human thioredoxins: evidence for a regulatory homodimer. Weichsel, A; Gasdaska, JR; Powis,
G; Montfort, WR. Structure 4, 735- 751, 1996.
2. Increasing the size of microcrystals by fine sampling of pH limits. A. McPherson. J. Appl. Cryst. (1995). 28, 362-365.
s t o c k o p t i o ns s o d i um a c e t a t e
O
ONa H
3
C
3H
2
O
pH 3.6 - 5.6
Order Information
Each StockOptions Citric Acid kit contains 44 unique reagents. To order individual reagents, use
Custom Shop

catalog number listed below. Refer to page 37 for further details.

Cat. No. Name Description Price
HR2-104 StockOptions Citric Acid 10 ml, tube format $195.00
HR2-904-** StockOptions Citric Acid Custom Shop 185 ml $144.00
** = reagent number 1-44
Order Information
Each StockOptions Sodium Acetate kit contains 21 unique reagents. To order individual reagents,
use Custom Shop

catalog number listed below. Refer to page 37 for further details.

Cat. No. Name Description Price
HR2-233 StockOptions Sodium Acetate 10 ml, tube format $195.00
HR2-933-** StockOptions Sodium Acetate Custom Shop 185 ml $150.00
** = reagent number 1-21
StockOptions Citric Acid buffer kit is a preformulated, sterile fltered set of titrated buffer stocks.
The reagents are supplied as 1.0 M stock solutions in 10 ml volumes. Each StockOptions Citric Acid
buffer reagent is carefully titrated using Sodium hydroxide. StockOptions Citric Acid is comprised of
44 unique reagents covering the pH range of 2.2 to 6.5 in 0.1 pH unit increments.
References
1. Increasing the size of microcrystals by ne sampling of pH limits. A. McPherson. J. Appl. Cryst. (1995). 28, 362-365.
s t o c k o p t i o ns c i t r i c a c i d
pH 2.2 - 6.5
OH O O
OH O
OH HO
d e s c r i p t i o n
StockOptions Buffer kits are reagent
toolboxes for the macromolecular crystal
grower. StockOptions reagent kits offer
precisely formulated, high quality crystal-
lization grade reagent stocks in convenient,
cost-effective kits. The chemicals utilized in
StockOptions kits are the same crystallization
grade, ultra-high purity chemicals utilized
in the Hampton Research kits. StockOptions
reagents are carefully formulated under strict
quality standards to ensure reliable performance and lot-to-lot consistency.
Preformulated stocks remove calculation, measurement, and formulation errors. No more second
guessing how the reagent was formulated, what specific chemical was used, when it was made, or how
to precisely reproduce that reagent when it is gone and more reagent is required for additional setups.
s T o C k o P T I o n s

B U F F E R
65
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Order Information
Each StockOptions Sodium Citrate kit contains 24 unique reagents. To order individual reagents, use
Custom Shop

catalog number listed below. Refer to page 37 for further details.

Cat. No. Name Description Price
HR2-235 StockOptions Sodium Citrate 10 ml, tube format $195.00
HR2-935-** StockOptions Sodium Citrate Custom Shop 185 ml $173.00
** = reagent number 1-24
StockOptions Sodium Citrate buffer kit is a preformulated, sterile filtered set of titrated buffer stocks.
The reagents are supplied as 1.0 M stock solutions in 10 ml volumes. Each StockOptions Sodium Citrate
buffer reagent is carefully titrated using Hydrochloric acid. The kit is comprised of 24 unique reagents
covering the pH range of 4.2 - 6.5 in 0.1 pH unit increments.
Formulated using Sodium citrate tribasic dihydrate and HCl.
pKa1 = 3.1 pKa2 = 4.8 pKa3 = 5.4
References
1. Increasing the size of microcrystals by fine sampling of pH limits. A. McPherson. J. Appl. Cryst. (1995). 28, 362-365.
s t o c k o p t i o ns s o d i um c i t r a t e b uf f e r
NaO
O
ONa
ONa
HO
H
2
O
H
2
O
O
O
CH
3
3H
2
O As
O
NaO CH
3
OH
S
O
O
O N
H
2
O
pH 4.2 - 6.5
StockOptions Sodium Cacodylate buffer kit is a preformulated, sterile filtered set of titrated buffer
stocks. The reagents are supplied as 1.0 M stock solutions in 10 ml volumes. Each StockOptions
Sodium Cacodylate buffer reagent is carefully titrated using Hydrochloric acid. The kit is comprised of
24 unique reagents covering the pH range of 5.1 to 7.4 in 0.1 pH unit increments.
References
1. Increasing the size of microcrystals by fine sampling of pH limits. A. McPherson. J. Appl. Cryst. (1995). 28, 362-365.
s t o c k o p t i o ns s o d i um c a c o d y l a t e b uf f e r
pH 5.1 - 7.4
StockOptions MES buffer kit is a preformulated, sterile filtered set of titrated buffer stocks. The
reagents are supplied as 1.0 M stock solutions in 10 ml volumes. Each StockOptions MES buffer reagent
is carefully titrated using Sodium hydroxide. The kit is comprised of 20 unique reagents covering the
pH range of 5.2 to 7.1 in 0.1 pH unit increments.
References
1. Characterization, crystallization and preliminary crystallographic analysis of human recombinant cyclooxygenase-2. Marco, SD; Priestle, JP; Grutter, MG;
Wennogle, LP; Boyar, W. Acta Crystallogr D 53, 224- 226, 1997.
2. Increasing the size of microcrystals by ne sampling of pH limits. A. McPherson. J. Appl. Cryst. (1995). 28, 362-365
s t o c k o p t i o ns me s b uf f e r
pH 5.2 - 7.1
Order Information
Each StockOptions Sodium Cacodylate kit contains 24 unique reagents. To order individual
reagents, use Custom Shop

catalog number listed below. Refer to page 37 for further details.

Cat. No. Name Description Price
HR2-239 StockOptions Sodium Cacodylate 10 ml, tube format $250.00
HR2-939-** StockOptions Sodium Cacodylate Custom Shop 185 ml $221.00
** = reagent number 1-24
Order Information
Each StockOptions MES kit contains 20 unique reagents. To order individual reagents, use Custom
Shop

catalog number listed below. Refer to page 37 for further details.

Cat. No. Name Description Price
HR2-243 StockOptions MES 10 ml, tube format $195.00
HR2-943-** StockOptions MES Custom Shop 185 ml $138.00
** = reagent number 1-20
O
O
S ONa HO
N N
pH 6.8 - 8.2
Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com 66
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StockOptions Bis-Tris buffer kit is a preformulated, sterile filtered set of titrated buffer stocks. The
reagents are supplied as 1.0 M stock solutions in 10 ml volumes. Each StockOptions Bis-Tris buffer
reagent is carefully titrated using Hydrochloric acid. The kid is comprised of 21 unique reagents cover-
ing the pH range of 5.5 to 7.5 in 0.1 pH unit increments.
References
1. Increasing the size of microcrystals by fine sampling of pH limits. A. McPherson. J. Appl. Cryst. (1995). 28, 362-365.
Each kit contains 15 unique 10 ml volumes of 1.0 M HEPES buffer solution titrated to the indicated
pH using NaOH.
References
1. The expression, purification and crystallization of the epsilon subunit of the F1 portion of the ATPase of Escherichia coli. Codd, R; Cox, GB; Guss, JM;
Solomon, RG; Webb, D. J Mol Biol 228, 306- 309, 1992.
2. Increasing the size of microcrystals by fine sampling of pH limits. A. McPherson. J. Appl. Cryst. (1995). 28, 362-365.
s t o c k o p t i o ns b i s - t r i s
s t o c k o p t i o ns he p e s
StockOptions Sodium HEPES buffer kit is a preformulated, sterile filtered set of titrated buffer stocks.
The reagents are supplied as 1.0 M stock solutions in 10 ml volumes. Each StockOptions Sodium HEPES
buffer reagent is carefully titrated using Hydrochloric acid. The kit is comprised of 15 unique reagents
covering the pH range of 6.8 to 8.2 in 0.1 pH unit increments.
References
1. Increasing the size of microcrystals by fine sampling of pH limits. A. McPherson. J. Appl. Cryst. (1995). 28, 362-365.
s t o c k o p t i o ns s o d i um he p e s
s T o C k o P T I o n s

B U F F E R
pH 6.8 - 8.2
pH 5.5 - 7.5
N
OH
OH
HO
HO
HO
S
O
O
OH HO
N N
Order Information
Each StockOptions Bis-Tris kit contains 21 unique reagents. To order individual reagents, use
Custom Shop

catalog number listed below. Refer to page 37 for further details.

Cat. No. Name Description Price
HR2-106 StockOptions Bis-Tris 10 ml, tube format $195.00
HR2-906-** StockOptions Bis-Tris Custom Shop 185 ml $151.00
** = reagent number 1-21
Order Information
Each StockOptions HEPES kit contains 15 unique reagents. To order individual reagents, use
Custom Shop

catalog number listed below. Refer to page 37 for further details.

Cat. No. Name Description Price
HR2-102 StockOptions HEPES 10 ml, tube format $195.00
HR2-902-** StockOptions HEPES Custom Shop 185 ml $172.00
** = reagent number 1-15
Order Information
Each StockOptions Sodium HEPES kit contains 15 unique reagents. To order individual reagents,
use Custom Shop

catalog number listed below. Refer to page 37 for further details.

Cat. No. Name Description Price
HR2-231 StockOptions Sodium HEPES 10 ml, tube format $195.00
HR2-931-** StockOptions Sodium HEPES Custom Shop 185 ml $190.00
** = reagent number 1-15
67
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NH
2
OH
OH HO
NH
2
OH
OH HO HCl
pH 7.0 - 9.0
pH 7.0 - 9.0
StockOptions Tris buffer kit is a preformulated, sterile filtered set of titrated buffer stocks. The reagents
are supplied as 1.0 M stock solutions in 10 ml volumes. Each StockOptions Tris buffer reagent is carefully
titrated using Hydrochloric acid. The kit is comprised of 21 unique reagents covering the pH range of 7.0
to 9.0 in 0.1 pH unit increments.
References
1. Crystallization and Preliminary X-ray Diffraction Analysis of the ArsC Protein from the Escherichia coli Arsenical Resistance Plasmid, R773. deMel,
VSJ; Doyle, MA; Gladysheva, TB; Oden, KL; Martin, PD; Rosen, BP; Edwards, BFP. J Mol Biol 242, 701- 702, 1994.
2. Increasing the size of microcrystals by fine sampling of pH limits. A. McPherson. J. Appl. Cryst. (1995). 28, 362-365.
StockOptions Tris Hydrochloride buffer kit is a preformulated, sterile filtered set of titrated buffer
stocks. The reagents are supplied as 1.0 M stock solutions in 10 ml volumes. Each StockOptions Tris
Hydrochloride buffer reagent is carefully titrated using Sodium hydroxide. The kit is comprised of 21
unique reagents covering the pH range of 7.0 to 9.0 in 0.1 pH unit increments.
References
1. Increasing the size of microcrystals by fine sampling of pH limits. A. McPherson. J. Appl. Cryst. (1995). 28, 362-365.
s t o c k o p t i o ns t r i s
s t o c k o p t i o ns t r i s hy d r o c hl o r i d e
"The Lord of the Ring" - Crystal of bacterial di-haem Cyt c4
grown using the Hampton Research Additive Screen.
Ivana Tomcova, Institute of Physical Biology,
Academy of Science of the Czech Republic.

Order Information
Each StockOptions Tris kit contains 21 unique reagents. To order individual reagents, use Custom
Shop

catalog number listed below. Refer to page 37 for further details.

Cat. No. Name Description Price
HR2-100 StockOptions Tris 10 ml, tube format $195.00
HR2-900-** StockOptions Tris Custom Shop 185 ml $169.00
** = reagent number 1-21
Order Information
Each StockOptions Tris Hydrochloride kit contains 21 unique reagents. To order individual reagents,
use Custom Shop

catalog number listed below. Refer to page 37 for further details.

Cat. No. Name Description Price
HR2-237 StockOptions Tris Hydrochloride 10 ml, tube format $195.00
HR2-937-** StockOptions Tris Hydrochloride Custom Shop 185 ml $139.00
** = reagent number 1-21
As in selecting reagents for crystallization, the selection
of a suitable cryoprotectant involves some trial and error
as well as a screening.
1-15
A suitable cryoprotectant, when
mixed with the crystal and crystallization reagent, will
cool to cryogenic temperature without ice formation and
damage to the crystal. To assay for the proper concentra-
tion of cryoprotectant in the reagent used to grow the
crystal, one can mix the cryoprotectant with the crystal-
lization reagent and employ the desired cooling method.
For example, place the solution in a CryoLoop and
place the CryoLoop in a cryostream. Observe for ice formation either visually or with x-ray diffraction.
Upon cooling, a transparent drop and x-ray diffraction pattern, free of powder diffraction rings or ice
rings indicates success. The appearance of a cloudy drop or ice rings, indicates an inappropriate
cryoprotectant concentration or cryoprotectant. Incrementally increase the concentration and/or com-
position of the cryoprotectant serially 5 to 10% and repeat until the cooled drop remains clear while in
the cryostream. Once a clear drop is achieved in the cryostream, this is typically a good starting point
for cryopreservation of the crystal.
Some crystals can simply be dipped or washed quickly in a simple cryoprotectant such as 30% glycerol
for successful cryopreservation. However, when all else fails, a rational assay of each cryoprotectant
with incremental increases in cryoprotectant concentration, as well as a test of mixtures (i.e. a mixture
of sugars or a sugar mixed with Ethylene glycol) may be required to determine the best cryoprotectant
for a crystal.
Each CryoPro - Cryoprotectant kit contains 36 unique cryoprotectants including organics, oils, polyols,
salts, sugars, and polymers. 35 of the 36 reagents are supplied in 10 ml volumes. A single reagent L-(+)-
2,3-Butanol is supplied in a 0.2 ml aliquot. CryoPro is convenient and cost-effective.
Ready-to-use cryoprotectants are sterile filtered and formulated with ultra-pure water, using high
purity reagents. Many of the individual cryoprotectants are available as a 100 ml or 200 ml Optimize
reagent. Please refer to the CryoPro Formulations pdf file to find the catalog number for the equivalent
Optimize.
References
1. Boutron, P. (1987). Non-equilibrium formation of ice in aqueous solutions: efficiency of polyalcohol solutions for vitrification. In: Pegg, D.E. & Karow,
A.M. Jr. (eds). The biophysics of organ prese
2. Garman, E.F., & Mitchell, E.P. (1996). Glycerol concentrations required for cryoprotection of 50 typical protein crystallization conditions. J. Appl. Cryst. 29,
584-587.
3. Garman, E.F., & Schneider, T.R. (1997). Macromolecular Cryocrystallography. J. Appl. Cryst. 30, 211-237.
4. Hope, H. (1988). Cryocrystallography of biological macromolecules: a generally applicable method. Acta Cryst. B 44, 22-26.
5. Kottke, T., & Stalke, D. (1993). Crystal handling at low temperatures. J. Appl. Cryst. 26, 615-619.
6. Kwong, P.D. Liu, Y. (1999). Use of cryoprotectants in combination with immiscible oils for flash cooling macromolecular crystals. J. Appl. Cryst. 32, 102-105.
7. Mehl, P. (1989). Experimental dissection of devitrification in aqueous solutions in 1,3-butanediol. Cryobiology. 26, 567-568.
8. Parkin, S., & Hope, H. (1998). Macromolecular cryocrystallography: Cooling, mounting, storage and transportation of crystals. J. Appl. Cryst. 31, 945-953.
9. Petcock, J.M., Wang, Y.-F., DuBois, G.C., Harrison, R.W., & Weber, I.T. (2001). Effects of different post-crystallization soaking conditions on the diffraction
of Mtcp1 crystals. Acta Cryst. D57, 763-
10. Petsko, G.A. (1975). J. Mol. Biol. 96, 381-392.
11. Rodgers, D.W. (1994). Cryocrystallography. Structure. 2, 1135-1140. 12. Schneider, T.R. (1997). Cryocrystallography of biological macromolecules.
Acta Physica Polonica A. 91, 739-744.
12. Teng, T.-Y. (1990). J. Appl. Cryst. 23, 387-391.
13. Walker, L.J., Moreno, P.O., Hope, H. (1998). Cryocrystallography: effect of cooling medium on sample cooling rate. J. Appl. Cryst. 31, 954-965.
14. Watenpaugh, K.D. (1991). Curr. Op. Struct. Biol. 1, 1012.
15. Schneider, T.R. (1997), Cryocrystallography of biological macromolecules. Acta Physica Polonica A. 91, 739-744.
n
Cryoprotectant reagent kit
n
36 unique cryoprotectants
n
Includes cryoprotectant tutorial
n
Preformulated, ready-to-use
n
Polyols, organics, oils, polymers, sugars,
& salts
s T o C k o P T I o n s

-
C R Y O P R O

Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com 68
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d e s c r i p t i o n
Order Information
Each CryoPro kit contains 36 unique reagents.
Cat. No. Name Description Price
HR2-132 CryoPro 10 ml, tube format $285.00
c ry o p r o
Cryoprotectant
F o r m u l a t i o n s
100% Glycerol
100% Ethylene glycol
100% Polyethylene glycol 200
100% Polyethylene glycol 400
80% v/v Polyethylene glycol 600
60% w/v Polyethylene glycol 4,000
50% w/v Polyvinylpyrrolidone K 15
100% (+/-)-2-Methyl-2,4-pentanediol
6.0 M 1,6-Hexanediol
100% 1,2-Propanediol
100% Paratone-N
100% Paraffin Oil
100% NVH Oil
100% Dimethyl sulfoxide (DMSO)
100% 2-Propanol
100% Ethanol
100% Methanol
70% w/v D-(+)-Sucrose
35% w/v meso-Erythritol
70% w/v Xylitol
15% w/v myo-Inositol
20% w/v D-(+)-Raffinose pentahydrate
50% w/v D-(+)-Trehalose dihydrate
70% w/v D-(+)-Glucose monohydrate
100% 2,3-Butanediol
100% L-(+)-2,3-Butanediol
5.0 M Lithium acetate dihydrate
10.0 M Lithium chloride
4.0 M Lithium formate monohydrate
8.0 M Lithium nitrate
2.0 M Lithium sulfate monohydrate
3.4 M Sodium malonate pH 7.0
3.5 M Magnesium acetate tetrahydrate
5.0 M Sodium chloride
7.0 M Sodium formate
7.0 M Sodium nitrate
Reagent
Number
1
2
3
4
5
6
7
8
9
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
32
33
34
35
36
Crystals shaped like a dragonfly wing.
Yi-Wei Change, Institute of Molecular Biology,
Academia Sinica, Taiwan.
"Butterfly"-crystal of hydroxynitrile lyase from
Prunus amygdalus grown with the Hampton
Research Grid Screen PEG/LiCl.
Andreas Giessauf at the Institute for Chemie,
Karl-Franzens-Universitt Graz, Austria.
Crystals of the FERM domain of Focal Adhesion Kinase.
The final precipitant was derived from an initial hit obtained
with condition 17 from Hampton Research Crystal Screen.
Derek F.J. Ceccarelli, Samuel Lunenfeld Research Institute,
Mount Sinai Hospital, Toronto, Ontario, Canada.
c r y s t a l l i z a t i o n p l a t e s ,
h a r d w a r e & a c c e s s o r i e s
Crystals of a frameshift promoting RNA pseudoknot from the coronavirus IBV, grown using the Hampton Research Natrix screen.
Simon Pennell, MRC National Institute for Medical Research, United Kingdom.
T A B L E O F C O N T E N T S
72 - 77 2 4 w e l l c r y s t a l l i z a t i o n p l a t e s
78 - 79 4 8 w e l l c r y s t a l l i z a t i o n p l a t e s
79 7 2 w e l l c r y s t a l l i z a t i o n p l a t e s
80 - 90 9 6 w e l l c r y s t a l l i z a t i o n p l a t e s
90 3 8 4 w e l l c r y s t a l l i z a t i o n p l a t e s
91 - 95 c o v e r s l i d e s & r e l a t e d t o o l s
95 g l a s s s i t t i n g d r o p r o d s

96 m i c r o - b r i d g e s

97 9 w e l l g l a s s p l a t e
97 - 98 s e a l a n t s , s e a l i n g g r e a s e & o i l s
99 - 101 s e a l i n g f i l m s , t a p e s , m a t s & c o v e r s
102 - 103 d i a l y s i s b u t t o n s

, m e m b r a n e s & a p p l i c a t o r s
104 g r a n a d a c r y s t a l l i z a t i o n b o x

& c a p i l l a r i e s
P A G E S
2 4 W E L L C R Y S T A L L I Z A T I O N P L A T E S
Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com 72
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d e s c r i p t i o n
C R Y S C H E M

P L A T E
The Cryschem Plate is a 24 well sitting drop plate that is sealed
with clear sealing tape. The plate is supplied with a raised cover so
the plate can also be sealed with vacuum grease and plain glass or
plastic cover slides (22 mm diameter). The stackable, 24 well plates
have distortion-free, flat bottom wells and a concave sitting drop
post in the center of the well for concentric equilibration of the
drop with the reagent. Each plate is individually sealed. The top
of the plate and each of the 24 wells is a single, flat plane for smooth and easy seal application. Well
spacing provides for a large seal area between wells to prevent cross contamination and evaporation.
Approximate size: 15.1 cm x 10.6 cm x 2.2 cm. Approximate well size: 1.6 cm x 1.5 cm. Typical fill
volume: 500 to 1,000 l. Well capacity: 1.5 ml. Maximum drop volume on post: 40 l. Recommended
seal: HR3-511 Crystal Clear Sealing Tape (1.88 inch x 43.7 yard roll, with cutter) or HR4-511 Crystal
Clear Sealing Tape (1.88 inch x 60 yard roll, without cutter).
The Cryschem 24-1 SBS Plate is a smaller, SBS footprint 24 well sitting drop microplate that is sealed
with tape or film. The plate is supplied with a raised cover so the plate can also be sealed with vacuum
grease and plain glass or plastic cover slides (18 mm diameter). The stackable, 24 well plates have
distortion-free, flat bottom wells and a concave sitting drop post in the center of the well for concentric
equilibration of the drop with the reagent. Each plate is individually sealed. The top of the plate and
each of the 24 wells is a single, flat plane for smooth and easy seal application. Well spacing provides
for a large seal area between wells to prevent cross contamination and evaporation. Approximate size:
127.8 mm x 85.5 mm x 14.4 mm. Typical fill volume: 500 to 700 l. Maximum drop volume on post: 12
l. Recommended seal: HR3-609 Crystal Clear Sealing Film (100 pack), HR4-521 ClearSeal Film (100
pack) or HR4-506 Crystal Clear Sealing Tape (3 inch x 55 yard roll).
References
1. Purification, crystallization and preliminary X-ray crystallographic analysis of ST1022, a putative member of the Lrp/AsnC family of transcriptional regulators
isolated from Sulfolobus tokodaii strain 7. Shigeyuki Yokoyama et al. Acta Cryst. (2007). F63, 964966
n
Sitting drop crystallization

n
Heavy atom soaks

n
Seeding
n
Optically clear, concave sitting drop post

n
Smooth, flat top for easy seal
n
Optically clear polystyrene
a p p l i c a t i o n s
f e a t u r e s
m a t e r i a l
Order Information
Cat. No. Name Description Price
HR3-158 Cryschem Plate 24 plate case $95.00
HR3-160 Cryschem Plate 100 plate case $350.00
HR1-002 Cryschem 24-1 SBS Plate 50 plate case $150.00
12 mm 18 mm 22 mm Crystal Clear ClearSeal 1.88" 3" AlumaSeal II
Plate # Wells Cover Slide Cover Slide Cover Slide Sealing Film Film Tape Tape Film
SEALS
24 X
24 X X X
24 X
24 X
24 X
24 X
24 X X X
48 X X X
48 X
96 X X X
96 X X X
96 X X X
96 X X X
96 X X X
96 X
Cryschem
Cryschem 24-1 SBS
VDX
VDXm
Greiner ComboPlate
Linbro

Intelli-Plate 24 Well
Intelli-Plate 48 Well
VDX48
Douglas Instruments
CrystalClear Strips
Corning

Greiner CrystalQuick
Intelli-Plate
MRC/Swissci
MaSTeRBLOCk

A
B
C
D
1 3 5 2 4 6
Top Side
A
B
C
D
1 3 5 2 4 6
Top Side
Top
Side
73
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Cover Slide (or Sealing Tape)
Well of VDX
Crystallization Plate
Reservoir Solution
Dialysis Button
Vacuum
Grease
n
Hanging Drop Crystallization
n
Dialysis Crystallization with Dialysis Buttons
Cover Slide (or Sealing Tape)
Crystallization Droplet
Well of VDX
or Linbro Plate
Reservoir Solution
Micro-Bridge
n
Sitting Drop Crystallization with Micro-Bridges or Glass Rods
d e s c r i p t i o n
The Intelli-Plate 24-4 is a 24 well sitting drop plate for crystallization
screening and optimization. It features 4 reagent wells along the
y-axis (A-D) and 6 reagent wells along the x-axis (1-6). The reagent
reservoir is typically filled with 250 l and is capable of holding
between 200 and 650 l. The 4 drop wells are arranged at the top
of each well and can hold up to 8 l (5 l for hydrophobic reagents
such as MPD).
The Hampton Research ClearSeal Film and Sealing Film Applicator are used to seal the Intelli-Plate
when a sitting drop experiment is performed. The advanced film is a transparent, polyolefin film with
clear, pressure-sensitive, silicone adhesive. The film is pre-cut to perfectly fit 96 well crystallization
plates and is complete with 1 cm wide, perforated end tabs which makes handling and positioning
of the film easy, without fear of fingerprints getting into the view field. The Sealing Film Applicator is
a small, hand-held applicator which should be used to create a consistent film seal across the entire
crystallization plate. The Intelli-Plate can also be sealed using one of the clear sealing tapes supplied
in a roll format.
The height of all Intelli-Plate 24-4 is 0.560 inches. The Art Robbins Instruments catalog number for this
plate is 102-0004-00.
n
Sitting drop crystallization

n
4 drop wells per reservoir

n
Optically clear polystyrene
n
alphanumeric labeled drop wells visible under
microscope
a p p l i c a t i o n
f e a t u r e s
I N T E L L I - P L A T E

2 4 - 4 W E L L
Order Information
Cat. No. Name Description Price
HR3-114 Intelli-Plate 24-4 well 40 plate case $200.00
Proteins crystals.
Kasumi Kobayashi, Taiji Nakae and Hiroyuki Akama.
The Kitasato Institute, Kanagawa, Japan.

Intelli-Plate 24-4 well
Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com
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2 4 W E L L C R Y S T A L L I Z A T I O N P L A T E S
Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com 74
d e s c r i p t i o n
V D X

P L A T E
24 well crystallization plate for hanging drop or sitting drop vapor
diffusion crystallization (when used with Micro-Bridges or Glass
Rods), or Dialysis crystallization (when used with Dialysis Buttons).
Stackable, optically clear plastic 24 well plates with raised covers (to
allow room for cover slides) and flat bottoms for exceptional optics.
Raised, wide rings around each reservoir (well) minimize cross
contamination and allow each well to be individually sealed with
22 mm diameter circle or square cover slides. Plates are individu-
ally wrapped and supplied without sealant. 15.0 cm x 10.8 cm plate
footprint is convenient for manual pipetting and well access while
at the same time compatible with some automated liquid handling
systems. Approximate dimensions: 15.0 cm x 10.6 cm x 2.2 cm.
Approximate well size: 1.7 cm x 1.6 cm. Typical well volume: 500 to
1,000 l. Well capacity: 3.5 ml.
n
Hanging drop crystallization

n
Sitting drop crystallization with Micro-Bridges

or Glass Sitting Drop Rods

n
Dialysis crystallization with Dialysis Buttons

n
Compatible with 22 mm diameter square
and circle cover slides

n
24 well plate with raised cover
n
extremely versatile and cost-effective
crystallization platform
n
Optically clear plastic
a p p l i c a t i o n s
f e a t u r e s
Order Information
Cat. No. Name Description Price
HR3-142 VDX Plate 24 plate case $65.00
HR3-140 VDX Plate 100 plate case $250.00
d e s c r i p t i o n
V D X

P L A T E W I T H S E A L A N T
24 well crystallization plate for hanging drop or sitting drop vapor
diffusion crystallization (when used with Micro-Bridges or Glass
Rods), or dialysis crystallization (when used with Dialysis Buttons).
Stackable, optically clear plastic 24 well plates with raised covers (to
allow room for cover slides) and flat bottoms for exceptional optics.
Raised, wide rings around each reservoir (well) minimize cross
contamination and allow each well to be individually sealed with 22
mm diameter circle or square cover slides. Plates are individually
wrapped and supplied with applied sealant. 15.0 cm x 10.8 cm plate
footprint is convenient for manual pipetting and well access while
at the same time compatible with some automated liquid handling
systems. Approximate dimensions: 15.0 cm x 10.6 cm x 2.2 cm.
Approximate well size: 1.7 cm x 1.6 cm. Typical well volume: 500 to
1,000 l. Well capacity: 3.5 ml.
n
Hanging drop crystallization

n
Sitting drop crystallization with Micro-Bridges

or Glass Sitting Drop Rods

n
Dialysis crystallization with Dialysis Buttons
n
Sealant applied to 24 wells, ready to seal
with cover slides

n
Compatible with 22 mm diameter circle
and square cover slides
n
24 well plate with raised cover
n
Optically clear plastic
n
extremely versatile and cost-effective
crystallization platform
a p p l i c a t i o n s
f e a t u r e s
Order Information
Cat. No. Name Description Price
HR3-172 VDX Plate with sealant 24 plate case $110.00
HR3-170 VDX Plate with sealant 100 plate case $400.00
75
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d e s c r i p t i o n
d e s c r i p t i o n
24 well plate for hanging drop vapor diffusion crystallization with a
microplate footprint. Stackable, optically clear plastic, 24 well plates
with raised covers (to allow room for cover slides) and flat bottoms
for exceptional optics. Raised, wide rings around each reservoir
(well) minimize cross contamination and allow each well to be indi-
vidually sealed with 18 mm diameter circle cover slides. Plates are
individually wrapped and supplied without sealant. The microplate
footprint is space-saving and compatible with automated liquid
handling systems. Approximate dimensions: 128 mm x 85 mm. Approximate well ID: 14.4 mm. Typical
well volume: 100 to 500 l.
24 well plate with sealant for hanging drop vapor diffusion
crystallization with a 128 mm x 85 mm microplate footprint.
Stackable, optically clear plastic, 24 well plates with raised
covers (to allow room for cover slides) and flat bottoms for
exceptional optics. Raised, wide rings around each reservoir
(well) minimize cross contamination and allow each well to
be individually sealed with 18 mm diameter circle cover slides.
Plates are individually wrapped and supplied with applied seal-
ant. The microplate footprint is space-saving and compatible
with automated liquid handling systems. Approximate dimen-
sions: 128 mm x 85 mm. Approximate well ID: 14.4 mm. Typical
well volume: 100 to 500 l.
n
Hanging drop crystallization
n
24 well plate with raised cover

n
Compatible with 18 mm diameter circle cover
slides
n
Optically clear plastic
n
128 mm x 85 mm microplate footprint
n
Hanging drop crystallization
n
24 well plate with raised cover

n
Sealant applied to 24 wells, ready to seal
with cover slides
n
Compatible with 18 mm diameter circle cover
slides
n
Optically clear plastic
n
128 mm x 85 mm microplate footprint
a p p l i c a t i o n
a p p l i c a t i o n
f e a t u r e s
f e a t u r e s
Order Information
Cat. No. Name Description Price
HR3-108 VDXm Plate without sealant 50 plate case $130.00
Hanging Drop Crystallization
Order Information
Cat. No. Name Description Price
HR3-306 VDXm Plate with sealant 50 plate case $210.00
Crystals of a nucleoporin complex.
James Partridge, Massachusetts Institute of Technology, USA.

V D X M

P L A T E
V D X M

P L A T E W I T H S E A L A N T
2 4 W E L L C R Y S T A L L I Z A T I O N P L A T E S
Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com
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76
d e s c r i p t i o n
The Greiner ComboPlate is a 24 well plate with a raised cover in an
SBS microplate footprint. The plate, supplied without sealant, can
be sealed with an 18 mm diameter cover slide and sealant for hang-
ing drop crystallization. When used with the Greiner CrystalBridge,
the ComboPlate can be used for sitting drop crystallization and
sealed with sealing film/tape or 18 mm diameter cover slide and
sealant. Dimensions: 127.76 mm x 85.48 mm x 18.8 mm. Well
depth: 16.28 mm. 3.3 ml well capacity. Typical fill volume: 0.5 - 1.0
ml. Supplied with cover. The Greiner CrystalBridge is a polystyrene
sitting drop pedestal with a concave drop well (45 l capacity) and
is designed specifically for the ComboPlate.
n
Hanging and sitting drop crystallization

n
24 reagent wells

n
Sitting drop when used with CrystalBridge
n
ComboPlate has flat, raised rings about
each well to minimize spills and cross
contamination
n
ComboPlate seals with 18 mm glass or
plastic cover slides
n
Slightly raised protection lid
n
Polystyrene
a p p l i c a t i o n
f e a t u r e s
m a t e r i a l s
G R E I N E R C O M B O P L A T E

A N D C R Y S T A L B R I D G E

Order Information
Cat. No. Name Description Price
HR3-200 24 Well ComboPlate with cover, Greiner 662150 24 plate case $61.00
HR3-150 CrystalBridge for ComboPlate, Greiner 662145 100 pack $78.00
HR3-154 CrystalBridge for ComboPlate, Greiner 662145 400 pack $276.00
d e s c r i p t i o n
The Linbro 24 well plate with cover is made of clear, rigid, polysty-
rene and used primarily for hanging drop vapor diffusion crystal-
lization. Sitting drop vapor diffusion may be performed when used
with Micro-Bridges or Glass Sitting Drop Rods. Plates supplied
without sealant. Compatible with 22 mm diameter circle and
square cover slides. Each plate is individually wrapped and sterile.
Wide, raised rings around each reservoir minimizes cross contami-
nation and allows each individual well to be sealed using vacuum
grease, DC 7 Release Compound or immersion oil and a siliconized or plain cover slide. Cover is not
raised but can be raised by placing wax or mounting clay in each of the four corners of the cover. The
stackable plates have distortion-free, flat-bottom wells of fine clarity. Wells are identified by lettered
rows A through D and numbered columns 1 through 6. Approximate dimensions: 15.0 cm x 10.8 cm
x 2.2 cm. Approximate well size: 1.7 cm x 1.6 cm. Typical well volume: 700 to 1,000 l. Individual well
capacity: 3.5 ml
References
1. Expression, purification and crystallization of 2-oxo-hept-4-ene-1,7-dioate hydratase (HpcG) from Escherichia coli C. T. Adachi, A. Izumi, D. Rea, S.-Y. Park,
J. R. H. Tame and D. I. Roper. Acta Cryst. (2006). F62, 1010-1012.
n
Hanging drop crystallization

n
Sitting drop crystallization with Micro-Bridges

or Glass Sitting Drop Rods

n
Dialysis crystallization with Dialysis Buttons

n
24 well plate with cover
n
Compatible with 22 mm diameter circle and
square cover slides
n
Optically clear plastic
a p p l i c a t i o n s
f e a t u r e s
Order Information
Cat. No. Name Description Price
HR3-110 Linbro Plate 50 plate case $302.00
HR3-112 Linbro Plate 100 plate case $592.00
L I N B R O

P L A T E
77
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d e s c r i p t i o n
The Modular VDX Plate is a standard 24 well VDX Plate with
individually removable wells. The modular format, consisting
of a VDX frame and removable wells creates a very flexible
plate format for screening and optimization of crystallization
experiments.
Preliminary crystallization screens can be set in the Modular
VDX Plate. Once the results are scored, the wells can be
removed and organized into separate frames for optimization
and observation. Preliminary crystallization screens typically
evaluate a diverse array of reagents, reagent concentrations, pH values, and other variables that can
influence sample solubility and crystallization. These crystallization variables, combined in a single well
can have a unique effect on the sample solubility and are useful for screening preliminary crystallization
conditions. The crystallization screen typically delivers a wide range of solubility results, presented in
some wells as clear drops, others as precipitate, phase separation, or crystals. Clear drops can indicate
the relative supersaturation of the sample in the drop as too low for crystallization, while precipitate
can indicate the relative supersaturation as too high. If the sample has a temperature dependent solu-
bility, the temperature of the experiment can be raised or lowered, and the effect of this change can
be evaluated based upon the appearance of the drop. For example, a clear drop at room temperature
moved to 10C might result in a change in sample solubility, hence precipitate, phase separation,
or even a crystal. A precipitate at room temperature, moved to 30C may again result in a change in
sample solubility, hence precipitate, phase separation, or a crystal. When a standard crystallization
plate with fixed wells is used for screening, the decision to move the entire plate to a different tem-
perature can be complicated since all the wells in the plate must be moved together. This may be less
than ideal when crystals appears in one of those wells, or there is a mix of clear, precipitated, and phase
sep drops where each well requires a different course of action. Each solubility result or class (clear,
precipitate, phase sep, etc.) might require a unique temperature evaluation. With the Modular VDX
Plate, individual wells can be removed from the frame, organized by crystallization score or solubility
into a separate frame or frames, and the effect of temperature evaluated on a more discrete basis. For
example, clear drops can be removed from the screen frame, complete with their respective equili-
brated reservoir solution, placed into a separate frame and that newly created module can be incubated
at a lower temperature. Drops with precipitate can be removed from the screen frame, organized
into a new module and be incubated at an elevated temperature to see if an increase in temperature
improves sample solubility and promotes crystallization.
Beyond screening, the Modular VDX Plate also offers enhanced plate flexibility and customization
during optimization of preliminary crystallization experiments. Optimization experiments can be per-
formed in Modular VDX frames, utilizing removable wells to reorganize the plate as the optimization
proceeds. More promising conditions can be grouped into a single frame, reducing the actual utilized
active screen space required. Less promising experiments can be archived and stored into other
Modular VDX frames.
Identification of the Modular VDX Plates is simple. Frames and well sides can be labeled using a variety
of color coded Tough-Tag labels.
The Modular VDX Plates are also handy for the organization of drops and wells containing crystals.
Wells containing crystals suitable for diffraction analysis can be combined, stored, and carefully trans-
ported in modules, avoiding the extra baggage of wells containing clear drops and precipitate.
Just like the original VDX Plate, the Modular VDX Plate is compatible with Micro-Bridges

, Glass Sitting
Drop Rods, as well as 22 mm circle and square cover slides.
References
1. A modular plate for the optimization of crystallization experiments. J. Appl. Cryst. (2002) 35, 509-510.
n
Sitting, hanging drop, and dialysis
crystallization

n
Modular wells for temperature studies
1

n
Removable wells

n
24 well plate with raised cover
n
Wells available with or without sealant
n
Use with 22 mm diameter circle and square
cover slides
n
Optically clear plastic
a p p l i c a t i o n s
f e a t u r e s
Order Information
Cat. No. Name Description Price
HR3-198 Modular VDX Frames 24 frames with covers $57.00
HR3-196 Modular VDX Wells no sealant, 576 wells $70.00
HR3-204 Modular VDX Well with sealant, 96 wells $25.00
V D X

P L A T E ( M O D U L A R )
4 8 W E L L C R Y S T A L L I Z A T I O N P L A T E S
Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com 78
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d e s c r i p t i o n
The Intelli-Plate 48-2 is a 48 well sitting drop plate for otpi-
mization. The Intelli-Plate 48-2 features 6 reagent wells along
the y-axis (A-F) and 8 reagent wells along the x-axis (1-8). The
reagent reservoir is typically filled with 200 l and is capable of
holding between 100 and 500 l. The optimization drop wells
are arranged at the top of each well. The left sample drop well
holds between 4 l or less while the elongated sample well holds
20 l or less.
The IntelliPlate 48-3 is a 48 well sitting drop plate for screening.
The Intelli-Plate 48-3 features 6 reagent wells along the y-axis
(A-F) and 8 reagent wells along the x-axis (1-8). The reagent
reservoir is typically filled with 200 l and is capable of holding
between 100 and 500 l. The screening drop wells are arranged
at the top of each well. The three sample drop wells each hold
between 4 l or less.
The Hampton Research ClearSeal Film and Sealing Film
Applicator are used to seal the Intelli-Plate when a sitting drop
experiment is performed. The advanced film is a transparent,
polyolefin film with clear, pressure-sensitive, silicone adhesive.
The film is pre-cut to perfectly fit 96 well crystallization plates and
is complete with 1 cm wide, perforated end tabs which makes
handling and positioning of the film easy, without fear of fingerprints getting into the view field. The
Sealing Film Applicator is a small, hand-held applicator which should be used to create a consistent
film seal across the entire crystallization plate. The Intelli-Plate can also be sealed using one of the clear
sealing tapes supplied in a roll format.
The height of all Intelli-Plates is 0.560 inches.
n
Sitting Drop Crystallization

n
SBS Footprint

n
Intelli-Plate 48-2 Drop Volume:
20 l & 4 l
Reservoir: 500 l
n
Intelli-Plate 48-3 Drop Volume: 4 l
Reservoir: 500 l
a p p l i c a t i o n
f e a t u r e s
Order Information
Cat. No. Name Description Price
HR8-150 Intelli-Plate 48-2 well 10 plate case $85.00
HR8-152 Intelli-Plate 48-2 well 40 plate case $300.00
HR8-154 Intelli-Plate 48-3 well 10 plate case $85.00
HR8-156 Intelli-Plate 48-3 well 40 plate case $300.00
Individual Side Well View
Drop Support
Reservoir
Drop Support
Reservoir
Individual Top Well View Intelli-Plate & Intelli-Plate 48-2
Drop Support
Individual Top Well View Intelli-Plate 48-3
Reservoir
HR8-152
HR8-156
I N T E L L I - P L A T E 4 8 - 2 A N D 4 8 - 3
Top Side Well
Top Side Well
Top Side Well
79
Hanging Drop Crystallization
d e s c r i p t i o n
d e s c r i p t i o n
48 well plate with sealant for hanging drop vapor diffusion crys-
tallization with a microplate footprint. Stackable, optically clear
plastic, 48 well plates with raised covers (to allow room for cover
slides)and flat bottoms for exceptional optics. Raised, wide rings
around each reservoir (well) minimize cross contamination and
allow each well to be individually sealed with 12 mm diameter
circle cover slides. Plates are individually wrapped and supplied
with applied sealant. The microplate footprint is space-saving and
compatible with automated liquid handling systems. Approximate
dimensions: 128 mm x 85 mm. Approximate well ID: 9 mm. Typical
well volume: 100 to 300 l.
The 72 Well Microbatch Plate consists of a 6 by 12 well layout.
Paraffin, Silicon, or Als Oil is poured into the trough area of the
plate, submerging all 72 wells under oil. Sample and reagent are
then pipetted into each of the wells. The plate is available as a
plasma-treated, hydrophilic version for screening or an untreated,
hydrophobic version for optimization. The hydrophilic version
gives better liquid handling with small volumes, while the hydro-
phobic version reduces crystal nucleation and helps to prevent the
crystals from sticking to the plastic. Plate dimensions: 83.3 mm x 58.0 mm x 10.0 mm. Conical well with
11 l drop volume. Flat bottom well diameter: 1.3 mm. Supplied with cover.
n
Hanging drop crystallization
n
Sealant applied to 48 wells, ready to seal
with cover slide

n
Compatible with 12 mm diameter circle cover
slides
n
Optically clear plastic
n
128 mm x 85 mm microplate footprint
n
Microbatch crystallization
n
Conical, flat bottom wells

n
Hydrophobic or hydrophilic versions
n
Compact size
n
Supplied with cover
n
Polystyrene
a p p l i c a t i o n
a p p l i c a t i o n
f e a t u r e s
f e a t u r e s
m a t e r i a l
Order Information
Cat. No. Name Description Price
HR3-120 72 Well Microbatch Plate, Greiner 654180 treated, hydrophylic - $10.00
10 plate case
HR3-122 72 Well Microbatch Plate, Greiner 654180 treated, hydrophylic - $106.00
100 plate case
HR3-086 72 Well Microbatch Plate, Greiner 654102 untreated, hydrophobic - $10.00
10 plate case
HR3-087 72 Well Microbatch Plate, Greiner 654102 untreated, hydrophobic - $106.00
100 plate case
7 2 W E L L C R Y S T A L L I Z A T I O N P L A T E S
V D X 4 8

P L A T E W I T H S E A L A N T
Order Information
Cat. No. Name Description Price
HR3-275 VDX48 Plate with sealant 50 plate case $335.00
M I C R O B A T C H 7 2 W E L L P L A T E ( G R E I N E R )
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9 6 W E L L C R Y S T A L L I Z A T I O N P L A T E S
Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com 80
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CrystalClear Strips 96 well sitting drop plates are suitable for
both automatic and manual crystallization. They are particularly
useful for screening, since they feature a space-saving microplate
footprint. Sample and reagent drops (50 nl to 4 l) are dispensed
on a shelf on one side of each well. 50 to 100 l of crystallization
reagent is placed in the well. The strip is sealed with clear sealing
tape. Losses by evaporation though the body of the strip and the
tape are around 0.25% of the reservoir per day.
CrystalClear D (Douglas Instruments CCLEAR-D/1-10)
The CrystalClear D Strips, with indent (HR3-128) have a single
depression for the sample and reagent drop on the shelf above
each reservoir. This prevents hydrophobic reagents from running
off the shelf into the reservoir. This version should be used for
reagents and screens that contain low molecular weight alcohols
and detergents. The spacing between strips is 9 mm (the regular
SBS spacing). However, the spacing between sample chambers
along a D strip is only 8.45 mm. Therefore, if you want to use a
multichannel pipette to fill either the reservoirs or the sample wells,
use a 6 or 12-channel pipette.
CrystalClear P (Douglas Instruments CCLEAR-P/1-10)
The CrystalClear P Strips, without indent (HR3-162) have a small, raised, circular platform on the shelf
where samples are placed. This provides improved viewing but is not suitable for solutions containing
hydrophobic reagents such as low molecular weight alcohols and detergents. However, drops below
0.2 l (200 nl) containing alcohol (and other hydrophobic reagents) will generally work with all plates.
The spacing between strips is 9 mm (the regular SBS spacing). However, the spacing between sample
chambers along a P strip is only 8.45 mm. Therefore, if you want to use a multichannel pipette to fill
either the reservoirs or the sample wells, use a 6 or 12-channel pipette.
CrystalClear Duo (Douglas Instruments CCLEAR-DUO/1-10)
The CrystalClear Duo Strips is a 96 well sitting drop vapor diffusion plate. The plate features SBS 9
mm spacing in both directions, which makes the plate suitable for use with all dispensing robots and
in all observation systems. The plate offers low volume reagent wells for vapor diffusion with two
depressions on the shelf for samples. A plate consists of a single frame and twelve strips, snapped into
position, secured in the frame. Each strip contains eight wells. Each of the eight wells contains a single
reagent well and two sample drop wells. Sample drop wells are flat. Frame is color coded green.
CrystalClear Strips are packaged as twelve removable 8 well strips per frame (for a total of 96 wells per
frame) with ten frames (plates) per case. CrystalClear Strips are sealed using clear sealing tape or film.
Approximate plate size: 12.8 cm x 8.6 cm.
n
Sitting drop vapor diffusion crystallization
n
96 well sitting drop plate

n
Concave or flat drop area

n
Modular frame and well design

n
One or two sample drop wells per
reagent well
n
Polystyrene

a p p l i c a t i o n
f e a t u r e s
m a t e r i a l
Order Information
Cat. No. Name Description Price
HR3-128 CrystalClear D Strips, with indent 10 plate case $169.00
HR3-162 CrystalClear P Strips, without indent 10 plate case $169.00
DI-043 CrystalClear Duo Strips 10 plate case $145.00
HR3-128 CrystalClear D Strips, with indent HR3-162 CrystalClear P Strips, without indent DI-043 CrystalClear Duo Strips
HR3-162
HR3-128
C R Y S T A L C L E A R S T R I P S

( D O U G L A S I N S T R U M E N T S )
81
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Corning

CrystalEX 96 Well Crystallization plates are designed

for sitting drop vapor diffusion, high throughput protein crystal-
lization. The plates feature 96 protein wells corresponding to 96
reagent wells. The maximum reagent well volume is 210 l with
a typical working volume of 25 to 150 l. The maximum sample
well volume is 10 l for the conical bottom and 7 l for the flat
bottom. The flat bottom (1.5 mm diameter) sample wells provide
better crystal viewing for automated imaging systems. The well
surfaces are treated to be hydrophilic for improved drop mixing.
Both versions are compatible with 8 and 12 channel manual pipetters, automated liquid handling work
stations, and imaging systems. Both versions are manufactured from an advanced polymer with high
resistance to commonly used protein crystallization reagents. The advanced polymer material provides
for low-background polarization and high optical clarity so protein crystals may be viewed under polar-
ized light with minimal background interference. Low water absorption of the polymer prevents loss of
protein drop volume. The plates can be sealed using clear sealing tape or film.
n
Sitting drop crystallization
n
96 well sitting drop plate
n
Protein well shape is either conical bottom
(3773) or flat bottom (3785)

n
Compatible with 8 & 12 channel pipetters

n
advanced optically clear polymer

a p p l i c a t i o n
f e a t u r e s
m a t e r i a l
Order Information
Cat. No. Name Description Price
HR3-273 CrystalEX 96 Well, Conical Bottom, Corning 3773 10 plate case $110.00
HR3-271 CrystalEX 96 Well, Conical Bottom, Corning 3773 50 plate case $500.00
HR3-113 CrystalEX 96 Well, Flat Bottom, Corning 3785 10 plate case $110.00
HR3-115 CrystalEX 96 Well, Flat Bottom, Corning 3785 50 plate case $500.00
Individual Top Well View
Drop Support
Reservoir
Concave, Round Bottom
Drop support
Reservoir
Conical, Flat Bottom
Corning 3773 Corning 3785
Protein well shape Conical bottom Flat bottom
Protein well volume 10 l 7 l
Protein well dimensions (top/bottom) 3 mm 3 mm/1.5 mm
Protein well depth 3.1 mm 3.1 mm
Number of reagent/protein wells 96/96 96/96
Reagent well volume 210 l 210 l
Recommended reagent working volume 25-150 l 25-150 l
C R Y S T A L E X

( C O R N I N G )
Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com 82
9 6 W E L L C R Y S T A L L I Z A T I O N P L A T E S
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The second generation of Corning

96 Well, sitting drop

format plates are built to SBS specifications, making them
well suited for high throughput crystallization and are fully
compatible with robotic equipment. The plates are avail-
able in several different configurations with varying drop
well shapes, plate materials, and number of drop wells
per reagent well. The basic plate design is one reagent
well flanked by one or three drop wells, with SBS stan-
dard spacing between the centers of adjacent well clusters. One may choose a plate with small (1 l),
medium (2 l), or large (4 l) drop well volumes. The choice of round, flat, or conical flat well shapes
are available. The PZero polymer provides for zero background polarization and is non-birefringent.
PZero plates are not treated. The COC polymer offers high chemical resistance. Both types of plastic
feature improved transparency. The reservoir numbers are embossed on each individual well for easy
identification. Drop well locations conform to SBS standards for robotic handling. The low-volume
reagent well saves on reagent costs. The plates can be sealed using clear sealing tape or film.
n
Sitting drop crystallization
n
96 well sitting drop plate
n
Round, flat, and conical flat bottom drop well
shapes

n
1, 2 or 4 microliter drop wells
n
1, 3 or 5 drop wells per reservoir
n
Hydrophilicity treated or untreated
n
PZero or COC plate material
a p p l i c a t i o n
f e a t u r e s
Order Information
Cat. No. Name Description Price
HR8-135 Corning 3556 4 l round drop well, 1 drop well, COC, $105.00
untreated - 10 plate case
HR8-134 Corning 3556 4 l round drop well, 1 drop well, COC, $490.00
untreated - 50 plate case
HR8-137 Corning 3551 4 l conical flat drop well, 1 drop well, COC, $105.00
treated - 10 plate case
HR8-136 Corning 3551 4 l conical flat drop well, 1 drop well, COC, $490.00
treated - 50 plate case
HR8-139 Corning 3552 2 l round drop well, 3 drop well, PZero $105.00
- 10 plate case
HR8-138 Corning 3552 2 l round drop well, 3 drop well, PZero $490.00
- 50 plate case
HR8-141 Corning 3553 2 l conical flat drop well, 3 drop well, PZero $105.00
- 10 plate case
HR8-140 Corning 3553 2 l conical flat drop well, 3 drop well, PZero $490.00
- 50 plate case
HR8-147 Corning 3550 1 l conical flat drop well, 3 drop well, PZero $105.00
- 10 plate case
HR8-146 Corning 3550 1 l conical flat drop well, 3 drop well, PZero $490.00
- 50 plate case
Corning 3554 CrystalEx
HR8-143/42
Corning 3552 CrystalEx
HR8-139/38
Corning 3556 CrystalEx
HR8-135/34
Corning 3555 CrystalEx
HR8-145/44
Corning 3550 CrystalEx
HR8-146/47
Corning 3557 CrystalEx
HR8-160/158
Corning 3551 CrystalEx
HR8-137/36
Corning 3553 CrystalEx
HR8-141/40
Corning 3554 CrystalEx
HR8-143/42
Corning 3552 CrystalEx
HR8-139/38
Corning 3556 CrystalEx
HR8-135/34
Corning 3555 CrystalEx
HR8-145/44
Corning 3550 CrystalEx
HR8-146/47
Corning 3557 CrystalEx
HR8-160/158
Corning 3551 CrystalEx
HR8-137/36
Corning 3553 CrystalEx
HR8-141/40
Corning 3554 CrystalEx
HR8-143/42
Corning 3552 CrystalEx
HR8-139/38
Corning 3556 CrystalEx
HR8-135/34
Corning 3555 CrystalEx
HR8-145/44
Corning 3550 CrystalEx
HR8-146/47
Corning 3557 CrystalEx
HR8-160/158
Corning 3551 CrystalEx
HR8-137/36
Corning 3553 CrystalEx
HR8-141/40
Corning 3554 CrystalEx
HR8-143/42
Corning 3552 CrystalEx
HR8-139/38
Corning 3556 CrystalEx
HR8-135/34
Corning 3555 CrystalEx
HR8-145/44
Corning 3550 CrystalEx
HR8-146/47
Corning 3557 CrystalEx
HR8-160/158
Corning 3551 CrystalEx
HR8-137/36
Corning 3553 CrystalEx
HR8-141/40
C R Y S T A L E X

S E C O N D G E N E R A T I O N ( C O R N I N G )
Corning 3554 CrystalEx
HR8-143/42
Corning 3552 CrystalEx
HR8-139/38
Corning 3556 CrystalEx
HR8-135/34
Corning 3555 CrystalEx
HR8-145/44
Corning 3550 CrystalEx
HR8-146/47
Corning 3557 CrystalEx
HR8-160/158
Corning 3551 CrystalEx
HR8-137/36
Corning 3553 CrystalEx
HR8-141/40
Top Side Well
83
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Sitting drop vapor diffusion crystallization
n
Hanging drop vapor diffusion crystallization
with CrystalDrop

Lid
n
96 well sitting drop plate
n
Round or flat bottom drop wells

n
Hanging drop with CrystalDrop

Lid

n
Plastic
a p p l i c a t i o n
f e a t u r e s
m a t e r i a l
Order Information
Cat. No. Name Description Price
HR3-192 CrystalQuick 96 Well, 4 l square drop well, $130.00
Greiner 609101 3 drop well - 10 plate case
HR3-190 CrystalQuick 96 Well, 4 l square drop well, $490.00
Greiner 609101 3 drop well - 40 plate case
HR3-094G CrystalQuick Plus 96 Well, 4 l square drop well, 3 drop well, $139.00
Greiner 609830 LBR, hydrophobic - 10 plate case
HR3-095G CrystalQuick Plus 96 Well, 4 l square drop well, 3 drop well, $528.00
Greiner 609830 LBR, hydrophobic - 40 plate case
HR8-148 CrystalQuick Plus 96 Well, 4 l square drop well, 3 drop well, $130.00
Greiner 609130 hydrophobic - 10 plate case
HR8-149 CrystalQuick Plus 96 Well, 4 l square drop well, 3 drop well, $490.00
Greiner 609130 hydrophobic - 40 plate case
HR3-088 CrystalQuick 96 Well, 4 l square drop well, 3 drop well, $139.00
Greiner 609801 LBR, hydrophobic - 10 plate case
HR3-089 CrystalQuick 96 Well, 4 l square drop well, 3 drop well, $528.00
Greiner 609801 LBR, hydrophobic - 40 plate case
HR3-302 CrystalQuick 96 Well, 4 l square drop well, $230.00
Greiner 609171 1 drop well, low profile - 20 plate case
HR3-304 CrystalQuick 96 Well, 4 l square drop well, 1 drop well, $830.00
Greiner 609171 low profile - 80 plate case
HR3-092G CrystalQuick Plus 96 Well, 4 l square drop well, 1 drop well, $266.00
Greiner 609180 hydrophobic, low profile - 20 plate case
HR3-093G CrystalQuick Plus 96 Well, 4 l square drop well, 1 drop well, $1,011.00
Greiner 609180 hydrophobic, low profile - 80 plate case
HR3-284 CrystalQuick 96 Well, 4 l square drop well, 1 drop well, $255.00
Greiner 609871 LBR, low profile - 20 plate case
HR3-285 CrystalQuick 96 Well, 4 l square drop well, 1 drop well, $880.00
Greiner 609871 LBR, low profile - 80 plate case
HR3-283 CrystalQuick 96 Well, 2 l round drop well, $130.00
Greiner 609120 3 drop well - 10 plate case
HR3-281 CrystalQuick 96 Well, 2 l round drop well, $490.00
Greiner 609120 3 drop well - 40 plate case
HR3-090 CrystalQuick 96 Well, 2 l round drop well, $139.00
Greiner 609820 3 drop well, LBR - 10 plate case
HR3-091 CrystalQuick 96 Well, 2 l round drop well, $528.00
Greiner 609820 3 drop well, LBR - 40 plate case
HR3-096 CrystalDrop Lid, Greiner 609150 2 flat wells, ungreased - 10 lid case $54.00
HR3-097 CrystalDrop Lid, Greiner 609150 2 flat wells, ungreased - 40 lid case $201.00
HR3-137 Greased CrystalDrop Lid, 2 flat wells, greased - 10 lid case $233.00
Greiner 609050
HR3-138 Greased CrystalDrop Lid, 2 flat wells, greased - 40 lid case $850.00
Greiner 609050
The Greiner CrystalQuick 96 well sitting drop format plates are
built to SBS specifications, making them well suited for high
throughput crystallization and are fully compatible with robotic
equipment. The low volume reagent well saves on reagent costs.
The plates are available in round or square drop well shapes,
polystyrene, LBR and hydrophobic plate materials, and a varying
number of drop wells per reagent well. The CrystalQuick is avail-
able in a cyclic polyolefin low birefringence (LBR) material for
polarized crystal imaging, exceptional transparency, high chemical
resistance, and low water absorption. The low profile CrystalQuick version reduces the amount of
space required for storage. In combination with the CrystalDrop Lid, the CrystalQuick plates enable
simultaneous experiments using both sitting drop and hanging drop methodologies. The plates can be
sealed using clear sealing tape or film.
C R Y S T A L Q U I C K

9 6 W E L L S I T T I N G D R O P P L A T E ( G R E I N E R )
Top Side
Top Side
Top Side
Top
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1 2 3 4 5 6 7 8 9 10 11 12
Top Side
9 6 W E L L C R Y S T A L L I Z A T I O N P L A T E S
Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com 84
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The Intelli-Plate 96-2 Original crystallization plate is an optically
clear plate for sitting drop vapor diffusion crystallization. It has
been built to SBS specifications with 8 vertical wells versus 12
horizontal wells. This plate features two locations for sample per
reservoir. The sample drop locations are located along the left side
of the reagent reservoir, along the Y-axis of the plate. The sample
wells are concave depressions on the ledge above the adjacent, flat
bottom reagent reservoir. One sample well is located above the
second sample well. The top sample well can hold 10 l or less
of sample. The lower sample well can hold 4 l or less of sample. The reagent reservoir is typically
filled with 70 l of reagent and is capable of holding up to 140 l of reagent. The sidewalls separating
adjacent wells or reservoirs are 0.9 and 1.0 mm thick in order to offer a larger area for sealing the plate
and separation of the reservoirs. The vertical wells are labeled A-H along the left side of the plate and
the horizontal wells are labeled 1-12 along the top of the plate. The Intelli-Plate 96-2 is quite rigid,
with virtually no torsional flex and is designed for either manual or automated pipetting. All wells have
standard 9 mm spacing to conform to SBS standards. The Art Robbins Instruments catalog number
for this plate is 102-0001-00.
The Intelli-Plate 96-3 well crystallization plate is an optically clear plate for sitting drop vapor diffusion
crystallization. It has been built to SBS specifications with 8 vertical wells versus 12 horizontal wells.
This plate features three identical locations for sample per reservoir. The sample drop locations are
located along the left side of the reagent reservoir, along the Y-axis of the plate. The sample wells are
concave depressions on the ledge above the adjacent, flat bottom reagent reservoir. Each identical
well features a round bottom for easy crystal harvesting. Each drop well can hold up to 1 l of sample.
The reagent reservoir is typically filled with 70 l of reagent and is capable of holding up to 140 l of
reagent. The sidewalls separating adjacent wells or reservoirs are 0.9 and 1.0 mm thick in order to offer
a larger area for sealing the plate and separation of the reservoirs. The vertical wells are labeled A-H
along the left side of the plate and the horizontal wells are labeled 1-12 along the top of the plate. All
wells have standard 9 mm spacing to conform to SBS standards.
The Intelli-Plate 96 Flat Shelf crystallization plate is an optically clear plate for sitting drop vapor dif-
fusion crystallization. It has been built to SBS specifications with 8 vertical wells versus 12 horizontal
wells. This plate features a single flat shelf for sample drops. The sample drop shelves are located along
the left side of the reagent reservoir, along the Y-axis of the plate. The reagent reservoir is typically
filled with 100 l of reagent and is capable of holding up to 300 l of reagent. Maximum drop volume
is 10 l for the upper sample well, & 4 l for the lower sample well. The sidewalls separating adjacent
wells or reservoirs are 0.9 and 1.0 mm thick in order to offer a larger area for sealing the plate and
separation of the reservoirs. The vertical wells are labeled A-H along the left side of the plate and the
horizontal wells are labeled 1-12 along the top of the plate. The Intelli-Plate 96 Flat Shelf is quite rigid,
with virtually no torsional flex and is designed for either manual or automated pipetting. All wells have
standard 9 mm spacing to conform to SBS standards.
The Intelli-Plate 96-2 LVR is a Low Volume Reservoir version of the original Intelli-Plate. The Art
Robbins Instruments catalog number for this plate is 102-0001-00. The plates can be sealed using clear
sealing tape or film. The height of all Intelli-Plates is 0.560 inches.
n
Sitting drop crystallization
n
96 reservoir wells with 2 or 3 corresponding
sample wells

n
96 well Intelli-Plate is compatible with 8 and
12 channel pipetters for manual set up

n
Footprint and well spacing meet microplate
industry standards for automation

n
Intelli-Plate 96-2 well plate features a
maximum fill volume of 140 l for the
reservoir well, 10 l for the upper sample
well, & 4 l for the lower sample well

n
Compatible with robotic equipment for
automation
n
Intelli-Plate Flat Shelf 96 well plate features
a maximum fill volume of 300 l for the reser-
voir well and a flat shelf for the sample drop
n
Intelli-Plate 96-3 plate features a maximum
fill volume of 140 l for the reservoir well,
1 l for the sample well
n
Custom bar coding available - please inquire
a p p l i c a t i o n
f e a t u r e s
Order Information
Cat. No. Name Description Price
HR3-297 Intelli-Plate 96-2 Original 10 plate case $90.00
HR3-299 Intelli-Plate 96-2 Original 40 plate case $330.00
HR3-183 Intelli-Plate 96-3 well 10 plate case $90.00
HR3-185 Intelli-Plate 96-3 well 40 plate case $330.00
HR8-171 Intelli-Plate 96 Flat Shelf 10 plate case $90.00
HR8-172 Intelli-Plate 96 Flat Shelf 40 plate case $330.00
HR3-143 Intelli-Plate 96-2 LVR 10 plate case $90.00
HR3-145 Intelli-Plate 96-2 LVR 40 plate case $330.00
Intelli-Plate
TM
96-3 well Intelli-Plate
TM
96-2 well
Intelli-Plate 96-2 well Intelli-Plate 96-3 well
I N T E L L I - P L A T E

( A R T R O B B I N S I N S T R U M E N T S )
Top Side
Top Side
Top Side
Top Side
Top Side
MRC 2 Well Crystallization Plate
85
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The MRC 2 Well Crystallization Plate manufacted by Swissci is a 2
drop chamber, 96 well crystallization plate for sitting drop vapor
diffusion. This plate is sometimes referred to as the Innovaplate
SD-2 crystallography plate.
The plate was developed at the MRC Laboratory of Molecular
Biology (Cambridge, United Kingdom) in collaboration with Dr.
Jan Lwe. Designed to meet the stringent requirements specified
by crystallographers, and made of proprietary new materials that
facilitate enhanced imaging, the MRC Crystallization Plate provides
easier access and removal of crystals, improved mixing of reagent
and sample, and reduced sample and reagent volumes.
Easy Crystal Retrieval
Raised wide wells make the crystal mounting especially easy.
Easy Viewing
The wells are a wide conical shape and have a lens effect for perfect illumination. The micro-numbering
ensures you will never get lost again when using the microscope. The optically superior polymer is UV
transmissible and may be used to differentiate between salt and protein crystals.
Better Sealing
Wide partition walls between the wells give plenty of area for sealing with tape or film. No central
bending occurs in this very robust structure.
Wide Range of Volumes
Typical volumes are 50 to 100 l of reservoir and 100 nl to 5 l drop size. The 192 optical wells offer
twice the number of experiments of experimental constructions.
SBS Standard
The plate is designed to the 96 well SBS standard for all common holders and external alphanumeric
identification. The MRC Crystallization Plate is suitable for centrifugation.
The plates can be sealed using clear sealing tape or film.
Innovaplate is a trademark of Innovadyne Technologies, Inc.
n
Sitting drop crystallization
n
Lens effect drop wells for enhanced optics

n
Drop volume: 100 nl to 5 l

n
Micro-numbering alongside drop volumes

n
Reservoir volume: 50 to 100 l

n
SBS format
n
Rigid plate structure
n
Drop well allows easier crystal harvesting
n
Wide partition walls between wells improve
sealing
n
Developed in conjunction with the MRC
Laboratory of Molecular Biology in
Cambridge, United kingdom
n
Two drop wells per reservoir
a p p l i c a t i o n
f e a t u r e s
Order Information
Cat. No. Name Description Price
HR3-082 MRC 2 Well Crystallization Plate 10 plate case $130.00
HR3-083 MRC 2 Well Crystallization Plate 40 plate case $490.00
M R C 2 W E L L C R Y S T A L L I Z A T I O N P L A T E ( S W I S S C I )
Protein crystal.
Bebiana Moura,
Institute for Molecular and Cell Biology (IBMC) at Oporto, Portugal.

A
B
A
B
1
1
2
2
MRC 2 Well Crystallization Plate
9 6 W E L L C R Y S T A L L I Z A T I O N P L A T E S
Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com 86
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The MRC 3 Well Crystallization Plate is presented in a 96-well plate
format that offers unique properties which make it ideal for both
nanoliter crystallization screening and microliter optimization alike.
Made from optically superior polymer (UVP) and with a new design
of the wells, the plate allows easy crystal viewing and retrieval.
Easy Crystal Retrieval
Raised wide wells make the crystal mounting especially easy.
Easy Viewing
The wells are a wide conical shape and have a lens effect for perfect
illumination. The micro-numbering ensures you will never get lost
again (visible by microscope). The optically superior polymer is UV
transmissible and may be used to differentiate between salt and
protein crystals. Grown crystals are easy to identify and to remove
from well due to a low-binding polymer. Plate with 3 wells for each
sample, better growing security with triplicates or the ability to use
well two and well three as mixing stations. Wells fill without micro-droplets jumping out due to static
effects. The profile allows for easier storage. Low volume buffer well enables savings on reagents.
Better Sealing
Wide partition walls between the wells give plenty of area for good sealing with tape. No central bend-
ing occurs in this very robust structure.
Wide Range of Volumes
Typical volumes are 50 l of reservior and 100 nl to 5 l drop size. The 288 optical wells offer three
times the number of experimental constructions.
SBS Standard
The plates are designed to the 96-well SBS standard for all common holders and external numbering
(A-H, 1-12) with corner location make the plate easy to use in a robotic sampler. The 3 well plate is
suitable for centrifugation.
The plates can be sealed using clear sealing tape or film.
n
Sitting drop crystallization
n
Lens effect drop wells for enhanced optics

n
Drop volume: 100 nL to 5 l

n
Micro-numbering alongside drop volumes

n
Reservoir volume: 50 l

n
SBS format
n
Drop well allows easier crystal harvesting
n
Three drop wells per reservoir
n
Low profile for easier storage
a p p l i c a t i o n
f e a t u r e s
Order Information
Cat. No. Name Description Price
HR3-123 MRC 3 Well Crystallization Plate 10 plate case $125.00
HR3-125 MRC 3 Well Crystallization Plate 40 plate case $470.00
M R C 3 W E L L C R Y S T A L L I Z A T I O N P L A T E ( S W I S S C I )
MRC 3 Well Crystallization Plate
3 circular drop wells and 1 square reservoir
d e s c r i p t i o n
The Greiner 780261 MASTERBLOCK is a 96 Deep Well
polypropylene block (plate). Each well has a fill volume of
1.2 ml. The MASTERBLOCK is useful for holding crystal-
lization screen reagents when using liquid handling auto-
mation or multi channel pipets. Each plate is sterile and
packaged individually. Round bottom well for complete
reagent recovery. ClearChoice virgin polypropylene
resin (autoclavable to 121C). SBS recommended dimen-
sions. Alphanumerically coded wells with chimney-style, round well top.
Seal with AlumaSeal II, Crystal Clear Sealing Film, Cap Mats, or Robolid. Plate Lid is useful to cover the
opened MASTERBLOCK when not in immediate use to minimize evaporation. MASTERBLOCK is also
great for thermal sealing due to the raised chimney about each well.
n
96 Deep Well plate for crystallization screen
reagents


n
Sterile, individually sealed plates
n
Polypropylene
n
Seal with alumaSeal II, Crystal Clear Sealing
Film, Cap Mats, or Robolid
n
SBS format
a p p l i c a t i o n
f e a t u r e s
Order Information
Cat. No. Name Description Price
HR3-105 MASTERBLOCK 96 Deep Well polypropylene plate 50 plate case $210.00
M A S T E R B L O C K

9 6 D E E P W E L L P L A T E ( G R E I N E R )
MRC 3 Well Crystallization Plate
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The MRC Under Oil 96 Well Crystallization Plate is designed for
microbatch crystallization.
The plate was developed at the MRC Laboratory of Molecular
Biology (Cambridge, United Kingdom) in collaboration with Dr.
Jan Lwe and colleagues.
It is a result of many years of experience in successful robotic high-
throughput crystallization and combines many of the features not
earlier available to the crystallographer.
The new MRC Under Oil 96 Well Crystallization Plate is designed for
microbatch crystallization using paraffin, silicon oil, or a mixture of
the two. Following the initial experiment, evaporation of the drop
through the oil allows for second crystallization stage, enabling
further crystal growth as a consequence of concentration. This is
different from other experiments in that the conditions are then extreme in nature and permit new
conditions to arise.
The new MRC Under Oil 96 Well Crystallization Plate offers unique properties that make it ideal for
both nanoliter crystallization screening and microliter optimization alike. Made from an optically supe-
rior polymer and with a new design of the wells, the plate allows easy crystal viewing and retrieval.
The advantages of the new MRC Under Oil 96 Well Crystallization Plate:
Easy Crystal Retrieval
Raised wide wells make the crystal mounting especially easy
Easy Viewing
The wells are wide conical and have a polished surface on both sides important for perfect illumina-
tion. The micro numbering ensures that you will never get lost again (visible by microscope). The
optically superior polymer is even UV transmissible and may be used to differentiate between salt
and crystals.
Better Sealing
Wide partition walls between the wells give plenty of area for good sealing with tape for the initial
experiments of microbatch. No central bending occurs in this very robust structure.
Recommended Volumes
Typical volumes validated for these plates are 20 l of oil with a shot through sample delivery of 100
to 500 nl. The 20 l volume of the individual wells gives the user a wide range of macromolecular
crystallization possibilities.
SBS Standard
The plates are designed to the 96 well SBS standard for all common holders ad external numbering
(A - H, 1 - 12) with corner location that make the plate easy to use in a robotic sampler. The plate can
also be centrifuged for better results. The unique MRC Under Oil 96 well Crystallization plate offers a
new way of microbatch crystallography. The 96 wells are optically perfect, designed to observe crystals
under a microscope.
Unique Polymer
The proprietary polymer is optically perfect - ultra low binding and guaranteed to have central drop
location in the well. There are no static effects and thus micro-droplet jumping is avoided.
d e s c r i p t i o n
n
Microbatch crystallization

n
easy crystal retrieval
n
easy viewing
n
Drop Volume 100 to 500 nl
n
Oil Volume 20 l
n
SBS Standard
n
Ultra low binding polymer, no static
a p p l i c a t i o n
f e a t u r e s
Order Information
Cat. No. Name Description Price
HR3-102 MRC Under Oil Crystallization Plate 10 plate case $120.00
HR3-104 MRC Under Oil Crystallization Plate 40 plate case $432.00
M R C U N D E R O I L 9 6 W E L L C R Y S T A L L I Z A T I O N P L A T E ( S W I S S C I )
A
1
A
2
B B
1 2
MRC Under Oil
Crystallization Plate
MRC Under Oil Crystallization Plate
9 6 W E L L C R Y S T A L L I Z A T I O N P L A T E S
Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com 88
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d e s c r i p t i o n
The Greiner 96 Well Imp@ct Plate for microbatch crystallization
has conical, flat bottom wells that promote central location of the
sample drop and ease crystal harvesting. The plate has a flat, opti-
cally clear bottom. A raised perimeter wall surrounds the plate to
contain Paraffin Oil, Als Oil, or Silicon Oil. The entire inner area
of the plate or each individual sample well may be filled with oil.
The plate features a double rim reservoir to hold water or reagent
to control and manipulate vapor diffusion from the microbatch
drops. Plate dimensions: 127.76 mm x 85.48 mm x 14.4 mm. Conical flat bottom well with 8 l drop
volume. Flat bottom well diameter: 1.33 mm.
Greiner 673170 plate is clear. Greiner 673096 plate is black with clear drop area.
Plates supplied without cover. Plate Lids HR3-084 or HR3-085 (Greiner 656190) are available sepa-
rately.
The Greiner 673101 96 Well Imp@ct Plate has a low profile format
with individual conical shaped sidewalls and a smooth, flat bot-
tom for enhanced microscopic investigation of crystals. Conical
well shape avoids spreading of the sample and reagent droplet
away from the center of the well. Typical well capacity for Paraffin,
Silicon, or Als Oil is 10 to 20 l per well. Vertical wells are labeled
A-H. Horizontal wells are labeled 1-12. Stackable with supplied
cover. Plate Dimensions: 127.76 mm x 85.48 mm x 8.0 mm. Conical
well with 19 l drop volume. Flat bottom well diameter: 2 mm.
n
Microbatch crystallization

n
Conical, flat bottom wells
n
Clear and black Clear

n
Double rim reservoir
n
Polystyrene
n
Microbatch crystallization

n
Conical, flat bottom wells
n
Supplied with cover
n
Microplate footprint
n
Polystyrene
a p p l i c a t i o n
a p p l i c a t i o n
f e a t u r e s
f e a t u r e s
m a t e r i a l
m a t e r i a l
Order Information
Cat. No. Name Description Price
HR3-098 96 Well Imp@ct Plate, Greiner 673170 with reservoir, no cover - 10 plate case $118.00
HR3-099 96 Well Imp@ct Plate, Greiner 673170 with reservoir, no cover - 40 plate case $444.00
HR3-100 96 Well Imp@ct Plate, Greiner 673096 with reservoir, no cover, black Clear $135.00
- 10 plate case
HR3-101 96 Well Imp@ct Plate, Greiner 673096 with reservoir, no cover, black Clear $482.00
- 40 plate case
HR3-084 Plate Lid, Greiner 656190 10 lid case $10.00
HR3-085 Plate Lid, Greiner 656190 40 lid case $35.00
Order Information
Cat. No. Name Description Price
HR3-295 96 Well Imp@ct Plate, Greiner 673101 with cover, without reservoir $98.00
- 10 plate case
HR3-293 96 Well Imp@ct Plate, Greiner 673101 with cover, without reservoir $350.00
- 40 plate case
I M P @ C T

P L A T E W I T H R E S E R V O I R ( G R E I N E R )
I M P @ C T P L A T E W I T H O U T R E S E R V O I R ( G R E I N E R )
89
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The 96 Well Microbatch Plate is an 8 by 12 well microplate with
cover. Paraffin, Silicon, or Als Oil is dispensed into each individual
well. Sample and reagent are then pipetted into the wells below
the oil. Individual well bottoms are U shaped to center the drop
under oil. Oil can be rapidly (between 3 to 20 seconds) and neatly
dispensed into the plate using V&P Scientific VP195B Multi-Spense
or V&P Scientific VP195B1-96. After sample and reagent are dis-
pensed under the oil using an 8 channel and single channel pipet,
the plates are then centrifuged at very low speed for 5 to 10 minutes to mix and center the drops. The
covered plates can be stored between 0-37C and viewed free of condensation as the covers can be
removed for viewing without drying the drops when using the microbatch method.
n
Microbatch crystallization

n
96 concave bottom wells
n
Microplate footprint
n
Supplied with cover
n
Polystyrene
a p p l i c a t i o n
f e a t u r e s
m a t e r i a l
Order Information
Cat. No. Name Description Price
HR3-265 96 Well Microbatch Plate, with cover 10 plate case $50.00
HR3-267 96 Well Microbatch Plate, with cover 50 plate case $235.00
M I C R O B A T C H 9 6 W E L L P L A T E
d e s c r i p t i o n
The Douglas Instruments Vapor Batch Plate is designed for
both microbatch and sitting drop vapor diffusion crystal-
lization. It has 96 wells in the central region of the plate and
several reservoirs around the outside of the plate. Oil is placed
in the central region followed by sample and reagent for a
microbatch experiment. When the outside reservoirs are filled
with reagent, they can be used to preserve microbatch crys-
tals by preventing the drops from drying out. The reservoirs
can also be used for common dehydrant, sitting drop vapor
diffusion experiments where up to 96 wells are equilibrated against a single reagent or dehydrant.
Preliminary experiments suggest this method may find more hits in screening experiments than the
conventional method of using the same solution in the reservoir and the drop. Wells are individually
labeled. The Vapor Batch Plate is available as a plasma-treated hydrophilic version for screening or an
untreated, hydrophobic version for optimization. The hydrophilic version gives better liquid handling
with small volumes, while the hydrophobic version reduces crystal nucleation and helps to prevent
the crystals from sticking to the plastic. Vapor Batch Plate Holders which provide a Linbro

, VDX,
or SBS footprint are available on a special order basis. Approximate plate dimensions: 81 mm x 55 mm
x 20 mm. Drop well depth: 1.8 mm. Typical drop volume: 1 to 9 l. Maximum drop volume: 20 l.
Recommended oil fill: 2.5 ml. Recommended reagent/dehydrant volume: 8 ml.
n
Sitting drop and microbatch crystallization
n
anaerobic crystallization

n
96 conical, flat bottom wells
n
Supplied with cover
n
Microbatch and vapor diffusion from the
same plate
n
Compact size
n
Hydrophobic or hydrophilic versions
n
Polystyrene
a p p l i c a t i o n s
f e a t u r e s
m a t e r i a l
Order Information
Cat. No. Name Description Price
DI-038 Vapor Batch Plate, treated, hydrophilic 10 plate case $66.00
DI-040 Vapor Batch Plate, treated, hydrophilic 80 plate case $475.00
DI-039 Vapor Batch Plate, untreated, hydrophobic 10 plate case $66.00
DI-041 Vapor Batch Plate, untreated, hydrophobic 80 plate case $475.00
V A P O R B A T C H 9 6 W E L L P L A T E ( D O U G L A S I N S T R U M E N T S )
Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com 90
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d e s c r i p t i o n
The Multichannel Pipetter Basin is a polypropylene trough
that allows for rapid reproduction of similar reagents or oils
throughout an entire 96 Well crystallization plate. Basin comes
with cover.
n
Pipet oil reservoir for microbatch
crystallization
n
Reagent reservoir for crystallization reagents
when setting Silver Bullets screens

n
Basin with cover
n
Compatible with 8 & 12-channel pipets
n
Polypropylene
a p p l i c a t i o n s
f e a t u r e s
m a t e r i a l
M U L T I C H A N N E L P I P E T T E R B A S I N
Order Information
Cat. No. Name Description Price
HR3-269 Multichannel Pipetter Basin each $7.00
9 6 W E L L C R Y S T A L L I Z A T I O N P L A T E S
3 8 4 W E L L C R Y S T A L L I Z A T I O N P L A T E S
d e s c r i p t i o n
The Corning

CrystalEX is designed for full automation in crys-

tal screening and built to meet industry standards for 384 well
microplate footprint and well locations. Features 192 reservoir
wells with a 105 l volume and 192 corresponding protein wells
with a maximum volume of 4 l. Typical reagent well volumes are
50 l and sample well volumes are 1 l. The reagent wells and sam-
ple wells are positioned to be compatible with multihead dispens-
ing equipment (up to 96 well heads). The plate is manufactured
from an advanced polymer with high resistance to commonly
used solvents, including acetone, acetic acid, butanone, ethanol,
iso-propanol, methanol, DMSO, nitric acid (65%), sulfuric acid
(40%), hydrochloric acid (36%), and ammonia solution (33%). The
advanced polymer features low background polarization and high optical clarity which allows crystals
to be viewed under polarized light with minimal background interference. The low water absorption of
the polymer prevents loss of protein drop volume. The plates can be sealed using Crystal Clear Sealing
Tape or ClearSeal Film. The plate is not treated.
References
1. Expanding screening space through the use of alternative reservoirs in vapor-diffusion experiments. Janet Newman. Acta Cryst. (2005). D61, 490-493.
n
Sitting drop crystallization

n
192 sample wells & 192 reagent wells
n
Flat bottom sample well
n
Low background polarization
n
SBS footprint
n
advanced polymer
n
Use the CrystaleX 384 plate to screen four
different reservoir solutions against the same
crystallization solution to expand screening
space (Newman 2005).
a p p l i c a t i o n
f e a t u r e s
m a t e r i a l
m e t h o d o l o g i e s
Order Information
Cat. No. Name Description Price
HR8-056 CrystalEX 384 Well Flat Bottom Plate 10 plate case $210.00
HR8-058 CrystalEX 384 Well Flat Bottom Plate 50 plate case $875.00
C R Y S T A L E X

3 8 4 W E L L F L A T B O T T O M P L A T E ( C O R N I N G )
Reservoir
Individual side well view
Reservoir
Individual top well view
Drop Support
Drop Support
Individual side well view Individual top well view
91
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C O V E R S L I D E S & R E L A T E D T O O L S
d e s c r i p t i o n
Siliconized glass cover slides allow a droplet to be suspended in
a position which provides near optimal conditions for vapor dif-
fusion with the surrounding reservoir solution. These siliconized
glass cover slides provide a consistent, high quality finish for
crystallization experiments. The hydrophobic surface produces a
drop which stands well and does not flatten on the glass. The
siliconized surface prevents the adhesion of crystals and precipitate
onto the glass surface. Use vacuum grease, DC Release Compound,
or immersion oil to seal cover slide to plate. Available in 12, 18 or
22 mm diameter circles and 22 mm diameter squares. Available in 0.22 mm or 0.96 mm glass thickness.
The 0.96 mm thick slides are virtually unbreakable during crystallization handling.
n
Hanging, sitting or sandwich drop
crystallization
n
Siliconized, hydrophobic glass surface
n
12 mm, 18 mm, & 22 mm glass diameter
n
0.22 mm & 0.96 mm glass thickness
n
Circle & square
a p p l i c a t i o n s
f e a t u r e s
Order Information
Cat. No. Name Description Price
HR3-278T Siliconized circle cover slides for Tecan robot 0.5 oz pack (~240 slides) $40.00
HR3-280T Siliconized circle cover slides for Tecan robot 5 oz case (~2,400 slides) $380.00
HR3-277 12 x 0.22 mm Siliconized circle cover slides 0.5 oz pack (~240 slides) $40.00
HR3-279 12 x 0.22 mm Siliconized circle cover slides 5 oz case (~2,400 slides) $380.00
HR8-088 12 x 0.96 mm Thick Siliconized circle cover slides 1 oz pack (~106 slides) $25.00
HR8-090 12 x 0.96 mm Thick Siliconized circle cover slides 10 oz case (~1,060 slides) $238.00
HR3-239 18 x 0.22 mm Siliconized circle cover slides 0.5 oz pack (~100 slides) $22.00
HR3-241 18 x 0.22 mm Siliconized circle cover slides 5 oz case (~1,000 slides) $210.00
HR3-515 18 x 0.96 mm Thick Siliconized circle cover slides 1 oz pack (~45 slides) $20.00
HR3-517 18 x 0.96 mm Thick Siliconized circle cover slides 10 oz case (~450 slides) $190.00
HR3-231 22 x 0.22 mm Siliconized circle cover slides 1 oz pack (~120 slides) $25.00
HR3-233 22 x 0.22 mm Siliconized circle cover slides 10 oz case (~1,200 slides) $238.00
HR3-247 22 x 0.96 mm Thick Siliconized circle cover slides 3 oz pack (~75 slides) $27.00
HR3-249 22 x 0.96 mm Thick Siliconized circle cover slides 30 oz case (~750 slides) $256.00
HR3-215 22 x 0.22 mm Siliconized square cover slides 1 oz pack (~100 slides) $22.00
HR3-217 22 x 0.22 mm Siliconized square cover slides 10 oz case (~1,000 slides) $210.00
HR3-223 22 x 0.96 mm Thick Siliconized square cover slides 4 oz pack (~75 slides) $25.00
HR3-225 22 x 0.96 mm Thick Siliconized square cover slides 40 oz case (~750 slides) $238.00
S I L I C O N I Z E D G L A S S C O V E R S L I D E S
Size Thickness Fits
12 VDX48
Size Thickness Fits
12 VDX48
Size Thickness Fits
18 VDXm
Size Thickness Fits
18 VDXm
Size Thickness Fits
22 VDX
Size Thickness Fits
22 VDX
Size Thickness Fits
22 VDX
Thickness Fits
VDX
Size
22
Size Thickness Fits
Flying eagles crystals. A mutant of AppA photoreceptor protein.
Vladimira Dragnea, Department of Biology,
Indiana University, Bloomington, Indiana, USA.
Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com 92
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C O V E R S L I D E S & R E L A T E D T O O L S
d e s c r i p t i o n
The Cover Slide Gizmo dispenses siliconized or plain slides
for convenient drop building during hanging drop vapor diffu-
sion crystallization. Load the cover slides into the dispenser. A
clear plastic cover or sealed dispenser keeps slides clean. Slide
the loaded dispenser along the platform and six cover slides
are dispensed onto the platform. A black background on the
platform makes for easy drop visualization as one builds the
drop(s). Use finger or forceps to place the completed cover slides onto the crystallization plate.
Each Cover Slide Gizmo includes a single cover so you only need to order HR8-166 if you lose the cover
for the HR8-162 or HR8-164. Manual operation, no power required.
n
Dispense circle or square cover slides for
hanging drop crystallization

n
Dispense siliconized or plain cover slides
n
Platform displays one to six cover slides
at a time for 18 and 22 mm slides
n
Platform displays one to twelve cover slides
at a time for 12 mm slides
n
available for 12, 18 or 22 mm diameter
cover slides for either original (0.22 mm)
or thick (0.96 mm) glass thickness
n
Black platform for slides allows one to see if
drops are clear or precipitated
a p p l i c a t i o n
f e a t u r e s
Order Information
Cat. No. Name Description Price
HR8-167 Cover Slide Gizmo Use for 12 x 0.22 mm circle cover slides $125.00
HR8-168 Cover Slide Gizmo Use for 12 x 0.96 mm circle cover slides $125.00
HR8-169 Cover Slide Gizmo Use for 18 x 0.22 mm circle cover slides $125.00
HR8-170 Cover Slide Gizmo Use for 18 x 0.96 mm circle cover slides $125.00
HR8-162 Cover Slide Gizmo Use for 22 x 0.22 mm circle cover slides $125.00
HR8-163 Cover Slide Gizmo Use for 22 x 0.96 mm circle cover slides $125.00
HR8-164 Cover Slide Gizmo Use for 22 x 0.22 mm square cover slides $125.00
HR8-165 Cover Slide Gizmo Use for 22 x 0.96 mm square cover slides $125.00
HR8-166 Replacement Cover Fits HR8-162 or HR8-164 $15.00
Square Cover Slide Gizmo
Circle Cover Slide Gizmo
G L A S S C O V E R S L I D E G I Z M O D I S P E N S E R
Protein crystal grown using the Hampton Research Crystal Screen.
Enrique Lemus Fuentes, Jefe de carrera de ing. en alimentos,
Universidad Tecnolgica de la Mixteca.
93
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Plain, non-siliconized, hydrophilic glass cover slides. Useful for
hanging drop, sitting drop, and sandwich drop vapor diffusion
crystallization methods. The hydrophilic surface creates a drop
which is more flat than a drop on a siliconized surface. This offers
enhanced imaging for very small drops. Use vacuum grease, DC
Release Compound, or immersion oil to seal cover slide to plate.
n
Hanging, sitting or sandwich drop
crystallization

n
Hydrophilic glass surface
n
12 mm, 18 mm, & 22 mm glass diameter

n
0.22 mm & 0.96 mm glass thickness
n
Circle & square
a p p l i c a t i o n s
f e a t u r e s
Order Information
Cat. No. Name Description Price
HR3-207T 12 mm Plain circle cover slides for Tecan robot 0.5 oz pack (~240 slides) $38.00
HR3-209T 12 mm Plain circle cover slides for Tecan robot 5 oz case (~2,400 slides) $361.00
HR3-235 18 x 0.22 mm Plain circle cover slides 0.5 oz pack (~100 slides) $20.00
HR3-237 18 x 0.22 mm Plain circle cover slides 5 oz case (~1,000 slides) $190.00
HR3-227 22 x 0.22 mm Plain circle cover slides 1 oz pack (~120 slides) $23.00
HR3-229 22 x 0.22 mm Plain circle cover slides 10 oz case (~1,200 slides) $219.00
HR3-243 22 x 0.96 mm Thick Plain circle cover slides 3 oz pack (~75 slides) $25.00
HR3-245 22 x 0.96 mm Thick Plain circle cover slides 30 oz case (~750 slides) $238.00
HR3-211 22 x 0.22 mm Plain square cover slides 1 oz pack (~100 slides) $20.00
HR3-213 22 x 0.22 mm Plain square cover slides 10 oz case (~1,000 slides) $190.00
HR3-219 22 x 0.96 mm Thick Plain square cover slides 4 oz pack (~75 slides) $23.00
HR3-221 22 x 0.96 mm Thick Plain square cover slides 40 oz case (~750 slides) $219.00
P L A I N G L A S S C O V E R S L I D E S
d e s c r i p t i o n
RNase-free, hydrophobic covers are lighter weight than glass.
Plastic cover slides are ready to use without washing or siliconiza-
tion. Hydrophobic working surfaces are protected with clean
release liners to prevent RNase contamination. The plastic cover
slides remain flat and will not curl, even at high temperatures.
Plastic cover slides do not chip, crack, or break and are dust-free.
Square cover slides are supplied protected in a sandwich polyeth-
ylene film. The circle cover slides are packed between fiber-free
paper slips. Use vacuum grease, DC Release Compound, or immer-
sion oil to seal cover slide to plate.
n
Hanging, sitting and sandwich drop
crystallization

n
0.22 mm thickness
n
RNase-free; handle with forceps to prevent
contamination
n
Compatible with most crystallization reagents
n
Siliconization is not required
n
18 mm circle, 22 mm circle,
or 22 mm square
a p p l i c a t i o n
f e a t u r e s
Order Information
Cat. No. Name Description Price
HR8-082 22 mm OptiClear Plastic circle cover slides 100 pack $42.00
HR8-084 22 mm OptiClear Plastic circle cover slides 1,000 pack $373.00
HR8-074 22 mm OptiClear Plastic square cover slides 100 pack $28.00
HR8-076 22 mm OptiClear Plastic square cover slides 1,000 pack $248.00
HR8-078 18 mm OptiClear Plastic circle cover slides 100 pack $42.00
HR8-080 18 mm OptiClear Plastic circle cover slides 1,000 pack $373.00
O P T I C L E A R

P L A S T I C C O V E R S L I D E S
Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com 94
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C O V E R S L I D E S & R E L A T E D T O O L S
d e s c r i p t i o n
d e s c r i p t i o n
d e s c r i p t i o n
PEN-VAC is a great tool for handling small, flat surfaced objects such
as plain and siliconized cover slides, as well as plastic slides. Simply
depress the side button, touch the tip to the object to be lifted and
release the button. PEN-VAC creates a totally self-contained vacuum
that lifts up to 50 grams for up to one minute. Less than an ounce,
the lightweight aluminum body fits in a pocket. No power supply
needed. The kit includes one PEN-VAC with 5 3/4" blue body, (3)
angled and (3) straight probes with (2) each 1/8, 1/4, and 3/8 diameter blue silicone vacuum cups.
It is packaged in a clear plastic storage case.
The Cover Slide Vacuum Gadget allows one to pick up a single
cover slide, invert the slide and gadget, rest the gadget on the
bench so the slide rests as if on a pedestal, pipet the drop onto the
slide, then use the gadget to place and release the slide onto the
crystallization plate. The Gadget is a hand held vacuum bulb with
a single Buna-N vacuum cup. Squeeze the bulb, press the vacuum
cup to the slide, ease the squeeze on the bulb to create a vacuum
and the slide is held in place at the end of the Gadget. When ready
to position and place the slide give the Gadget a gentle squeeze
to release the vacuum. Single bulb with 3/8" (9.53 mm) diameter
cup.
A convenient holder for a crystallization plate, tilted at a 30 angle
to comfortably view samples and pipet without strain. Equipped
with a neoprene pad and rubber feet to prevent slippage. Works
well with smaller, 96 well footprint plates.
n
Holding and manipulating small, flat surfaces
such as glass cover slides

n
Self-contained vacuum
n
No power needed
n
Compact
n
Holding and manipulating small,
flat surfaces such as glass cover slides

n
Self-contained vacuum
n
No power needed
n
Compact
n
Convenient plate holder
a p p l i c a t i o n
a p p l i c a t i o n
a p p l i c a t i o n
f e a t u r e s
f e a t u r e s
Order Information
Cat. No. Name Description Price
HR3-251 PEN-VAC Kit each $95.00
Order Information
Cat. No. Name Description Price
HR8-098 Cover Slide Vacuum Gadget each $27.00
P E N - V A C

C O V E R S L I D E V A C U U M G A D G E T

P L A T E S T A N D
Order Information
Cat. No. Name Description Price
HR4-468 Plate Stand each $41.00
d e s c r i p t i o n
Solid Glass Sitting Drop Rods with a concave depression in the top
can hold up to 50 l and fit into the well of a 24 well VDX Plate
or Linbro

Plate. The rods can stand freely in the well, or a small

amount of vacuum grease can be applied to the bottom of the rod
to secure it to the plate. They also offer enhanced temperature
stability due to the mass of the rod. The rods are 14 mm tall, 10
mm in diameter, and the depth of the concave depression is 2 mm.
The rods are supplied without siliconization.
n
Sitting drop crystallization
n
Perform sitting drop experiments in 24 well
hanging drop plates
n
Made of glass, may be siliconized
n
Thermal mass stabilizes drop temperature
n
Optically clear glass
a p p l i c a t i o n
Order Information
Cat. No. Name Description Price
HR3-146 Glass Sitting Drop Rods 24 pack $100.00
HR3-148 Glass Sitting Drop Rods 144 pack $540.00
f e a t u r e s
m a t e r i a l
ATPase-domain complex of HlyB.
Jelena Zaitseva and Lutz Schmitt, Institute of Biochemistry,
Heinrich Heine University, Duesseldorf, Germany.
95
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Remove dust from cover slides, plates, and other laboratory
supplies.
n
Dust removal
a p p l i c a t i o n
Order Information
Cat. No. Name Description Price
HR4-411 Duster - Canned Air 10 oz can $15.00
D U S T E R - C A N N E D A I R
G L A S S S I T T I N G D R O P R O D S

M I C R O - B R I D G E S

Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com 96
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d e s c r i p t i o n
d e s c r i p t i o n
Micro-Bridges are small devices in the shape of a bridge that are
designed to carry out sitting drop vapor diffusion crystallization
when placed in a VDX Plate or Linbro

Plate. A single Micro-

Bridge fits neatly into the reservoir of a standard 24 well VDX Plate
for a sitting drop crystallization experiment. Once placed inside
the wells, Micro-Bridges are stable and there is no need to stick
them to the wells with grease or adhesive. It is therefore possible
to transfer them to other wells during or after a crystallization
experiment. Why sitting drop? Placing the droplet in the indentation greatly reduces the risk of losing
the protein by accident. Crystallization can be carried out in the presence of detergents and organic
solvents which are compatible with polystyrene (such as MPD, iso-propanol, and ethanol). The protein
drop is less affected by condensation problems. Soaking and seeding experiments can be carried out
easily. Crystals can be transported more securely. Larger drop volumes can be used. Micro-Bridges
have a concave indentation in the top surface of the bridge which holds the sample droplet during
a crystallization experiment and prevents the droplet from spreading over a large area. Made from
polystyrene, these parts are highly transparent and suitable for most crystallizations. The surface of the
indentation is highly polished to facilitate the visual inspection of the drops under a microscope. The
maximum drop volume for the Micro-Bridge is 35 l. Reservoirs can be sealed with plain cover slides
and vacuum grease or clear sealing tape.
Same great Micro-Bridges, but manufactured from clarified poly-
propylene. These Micro-Bridges are resistant to most organic
solvents, and are especially useful with crystallization experiments
that involve detergents and other hydrophobic reagents. They
resist drop-spreading typically observed when using reagents such
as detergents, volatile organics such as iso-propanol and ethanol,
and non-volatile organics such as MPD. These Micro-Bridges also
resist acetone, dioxane, acetonitrile, 2,2,2 trifluoroethanol, and
other aggressive organic solvents.
n
Sitting drop crystallization
n
Heavy atom soaks
n
Seeding
n
Perform sitting drop experiments in 24 well
hanging drop plates
n
Removable
n
Optically clear polystyrene
n
Sitting drop crystallization
n
Small molecule crystallization
n
Heavy atom soaks
n
Perform sitting drop experiments in 24 well
hanging drop plates
n
Removable
n
Clarified polypropylene
a p p l i c a t i o n s
a p p l i c a t i o n s
f e a t u r e s
f e a t u r e s
m a t e r i a l
m a t e r i a l
Cover Slide (or Sealing Tape)
Crystallization Droplet
Well of VDX
or Linbro Plate
Reservoir Solution
Micro-Bridge
Order Information
Cat. No. Name Description Price
HR3-310 Micro-Bridges 100 pack $65.00
HR3-312 Micro-Bridges 400 pack $361.00
Order Information
Cat. No. Name Description Price
HR3-340 Micro-Bridges Polypropylene 100 pack $75.00
HR3-342 Micro-Bridges Polypropylene 400 pack $255.00
M I C R O - B R I D G E S

M I C R O - B R I D G E S

P O L Y P R O P Y L E N E
9 W E L L G L A S S P L A T E
S E A L A N T S , S E A L I N G G R E A S E & O I L S
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When setting up a Sandwich Box, one pipets 25 ml of crystallization
reagent or dehydrant into the bottom of the Sandwich Box, then
places a support into the box. The 9 well depression plate is placed
upon the support. Drops of sample and reagent are pipetted into
the depressions and the lid of the Sandwich Box is sealed closed
with vacuum grease. Why Sandwich Boxes? They allow one to use
very, very large drops. They are optically superior to plastic plates
and glass slides, and offer different equilibration kinetics than other
crystallization plates. Each of the Sandwich Box components can be washed and reused so there is
little waste with the Sandwich Box Setup. These plates can be used during screening and optimiza-
tion, but are best suited for final optimization and production of crystals. Since the glass plates can be
removed from the boxes, crystal mounting is convenient. The siliconized 9 well depression plates are
also useful for heavy atom soaks, cryo solution dilution and transfers, and seeding experiments where
serial dilutions are involved. Sandwich Box Setups are available as a complete set or as individual
components so one can customize the system to meet their needs. Square cover slides can be used
to seal individual reservoirs but are not typically used in a crystallization setup. The Sandwich Box
Setup consists of a square plastic box (4 5/16 x 4 5/16 x 1 1/8), a polystyrene plate support, and a
siliconized, glass plate (4 L x 3 3/8 W (100 mm x 85 mm)) with nine concave depressions (7/8 O.D.
x 1/4 D (22 mm x 7 mm)).
References
1. In situ X-ray crystallography. A. McPherson. J. Appl. Cryst. (2000). 33, 397-400.
n
Sitting drop crystallization
n
Seeding
n
Heavy atom soaks
n
Cryo transfers
n
Siliconized, 9 well glass plates
n
Concave wells
n
Polystyrene box and support
n
Siliconized glass and polystyrene
a p p l i c a t i o n s
Order Information
Cat. No. Name Description Price
HR3-136 Sandwich Box Setup contains 6 siliconized, 9 well glass plates, $299.00
6 plastic supports, and 6 sandwich boxes
with covers
HR3-134 Siliconized 9 Well Glass Plate 6 plate pack $273.00
HR3-132 Sandwich Box with cover 40 box case $93.00
f e a t u r e s
m a t e r i a l s
9 W E L L G L A S S P L A T E & S A N D W I C H B O X S E T U P
d e s c r i p t i o n
I M M E R S I O N O I L ( T Y P E A , B A N D N V H )
Useful for sealing cover slides to crystallization plates. Type A oil
has low viscosity (150 centistokes), Type B has medium viscosity
(1250 centistokes), Type 300 has medium/low viscosity (300 cen-
tistokes), and Type NVH has a very high viscosity (21,000 centist-
okes). This works well for incubations above 25C.
n
Useful for sealing cover slides to
crystallization plates
n
Cryoprotection
a p p l i c a t i o n s
Order Information
Cat. No. Name Description Price
HR3-611 Sample Pack Contains the following: Type a - 14.8 ml, Type B - 7.4 ml, $16.00
Type 300 - 7.4 ml, Type NVH - 7.4 ml
HR3-613 Type A 4 oz bottle $16.00
HR3-615 Type B 4 oz bottle $16.00
HR3-617 Type NVH 4 oz bottle $18.00
S E A L A N T S , S E A L I N G G R E A S E & O I L S
Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com
98
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The Dow Corning

7 Release Compound Grease sealant is similar

to vacuum grease but is less viscous. Being less viscous, it is easier
to push through a syringe when applying the sealant manually.
Squeeze the grease into a 10 cc syringe fitted with a 200 l pipet tip
trimmed to desired diameter and you are ready to grease.
D O W C O R N I N G

7 R E L E A S E C O M P O U N D G R E A S E
Order Information
Cat. No. Name Description Price
HR3-508 Dow Corning 7 Release Compound Grease 150 g tube $22.00
d e s c r i p t i o n
R E P L A C E M E N T G R E A S E C A R T R I D G E S
Replacement grease cartridges for the Grease Machine L-100
n
For the Grease Machine L-100
a p p l i c a t i o n
Order Information
Cat. No. Name Description Price
HR3-202 Grease Cartridge 5 pack $56.00
d e s c r i p t i o n
Dow Corning

Vacuum Grease. The standard sealant for hang-

ing, sitting, and sandwich drop vapor diffusion setups. Stiff, non-
melting, non-drying silicone sealant maintains its consistency from
-40 to 260C. Chemical-resistant and non-carbonaceous. Squeeze
the grease into a 10 cc syringe fitted with a 200 l pipet tip trimmed
to desired diameter and you are ready to grease, or try the Grease
Kit.
n
Standard sealant for hanging, sitting,
and sandwich drop vapor diffusion setups
a p p l i c a t i o n
Order Information
Cat. No. Name Description Price
HR3-510 Dow Corning Vacuum Grease 150 g tube $34.00
D O W C O R N I N G

V A C U U M G R E A S E
S E A L I N G F I L M S , T A P E S , M A T S & C O V E R S
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C R Y S T A L C L E A R S E A L I N G F I L M
Crystal Clear Sealing Film is an optically transparent sealing film
for SBS format 24, 48, 96 and 384 well plates. The 2 mil (0.05
mm) thick, optically clear, non-fluorescing polyester film with 1
mil (0.025 mm) thick acrylic custom adhesive is designed for the
convenient and secure sealing of crystallization plates. A split,
optically opaque polyester backing with two end tabs assures a
clean, uniform, adhesive layer and allows for easy and accurate
positioning on the plate. Before use, peel off the opaque center
protective polyester backing to reveal the optically clear sealing
film. Film length: 5.625" Film width: 3.125"; End tabs: 0.375"; Perforations: 0.415" from edge of film;
Corner radius: 3/32". DNase-, RNase-, and nucleic-acid-free, non-sterile. Recommended temperatures
from -40 to +120C.
n
Sealing film used to seal sitting drop
crystallization experiments
n
Fits SBS format microplates
n
No special applicator required
n
Optically clear
a p p l i c a t i o n
Order Information
Cat. No. Name Description Price
HR3-609 Crystal Clear Sealing Film 100 pack $108.00
f e a t u r e s
d e s c r i p t i o n
ClearSeal Film fits all SBS format 96 well plates. Minimal evapora-
tion, minimal cross-well contamination, easy to handle, and opti-
cally clear. Single coated, 2 mil (0.05 mm), clear polyolefin film with
pressure-sensitive silicone-based adhesive. Supplied as a 141.3 mm
x 79.4 mm film with polyester release liner (backing). Seals well to
polypropylene, polystyrene, and polycarbonate plates. Compatible
with aqueous and organic solvents. Suitable for use between -70
and 110C. For best seal, the plate must be sealed with the Sealing
Film Applicator. Limited shelf life. Must be used before expiration
date.
The Sealing Film Applicator is a specially designed tool for the
proper application of the ClearSeal Film. The design and rigid plas-
tic allows for the application of even and consistent pressure which
releases the pressure sensitive adhesive on the film to properly seal
the film to the plate.
n
Sealing film used to seal sitting drop
crystallization experiments
n
Optically clear
n
easy to handle
n
Pressure sensitive adhesive
n
For best results use specially designed
applicator
a p p l i c a t i o n
f e a t u r e s
C L E A R S E A L F I L M

A N D A P P L I C A T O R
Order information
Cat. No. Name Description Price
HR4-523 ClearSeal Film 25 pack $56.00
HR4-521 ClearSeal Film 100 pack $198.00
HR4-525 Sealing Film Applicator each $5.00
S E A L I N G F I L M S , T A P E S , M A T S & C O V E R S
Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com 100
d e s c r i p t i o n
The new revolutionary Swissci AG UVP Hanging Drop Crystallization
Plate Seal comes ready-to-use with dust free protective coating and
a specialty polymer base. The seals accommodate up to 3 separate
drops of protein and fit the standard 2 or 3 drop MRC plates. The
product has a 100 micron thin layer of UVP specialty polymer devel-
oped for compact drop creation and ability to shoot x-ray without
any noticeable diffraction.
The seals can be run with drops in the 96, 192 or 288 drop positions. The seal covers sit directly over
the microplate crystallization reagent wells and are closed with a long-term resistant closing adhesive
which is validated not to ingress into sample, nor lead to seal corruption. To remove an individual
crystal, the seal may be peeled back or simply cut out with a scalpel. The material is very thin, although
still remaining as a perfect evaporation barrier and can be sliced with a knife very easily.
The plate seals are produced in a class 10,000 clean room environment and guaranteed to be dust and
scratch free. Optical quality is of the highest level available. The seals are able to be used in the UV
range thus enabling the user to recognize salt from protein crystals under the ultra violet light source
microscope.
n
Hanging drop sealing film for MRC plates
n
Designed for use with the 2 or 3 drop MRC
plates
n
UV compatible
n
X-ray diffraction capability
a p p l i c a t i o n
f e a t u r e s
Order Information
Cat. No. Name Description Price
HR3-607 UVP Hanging Drop MRC Plate Seal 50 pack $470.00
U V P H A N G I N G D R O P M R C P L A T E S E A L
d e s c r i p t i o n
These optically clear tapes are compatible with protein crystal-
lization reagents. The Crystal Clear Sealing Tapes use solvent
based adhesives which are compatible with aqueous crystallization
reagents.
Catalog number HR3-511 is a 1.88 inch (48 mm) wide, 3 mil tape
on a 43.7 yard (40 M) roll with a 1.5 inch core and is supplied with
a green dispenser/cutter.
Catalog number HR4-511 is a 1.88 inch wide, 3 mil tape on a 60 yard
roll with a 3 inch core and no dispenser/cutter.
Two strips of the HR3-511 or HR4-511 will seal a Corning

, Cryschem, CrystalClear Strip, Greiner,

Intelli-Plate, Linbro

, MRC, VDX and VDXm plate.

Catalog number HR4-506 is a 3 inch wide, 3 mil tape on a 54.86 yard roll with a 3 inch core and no
dispenser/cutter.
One strip of the HR4-506 will seal a Corning, Cryschem 24-1 SBS, CrystalClear Strip, Greiner, Intelli-
Plate, MRC and VDXm plate.
See chart for tape and plate compatibility.
n
Sealing tape used to seal sitting drop
crystallization experiments
n
Optically clear
n
Compatible with a wide range of
crystallization reagents
a p p l i c a t i o n
Order Information
Cat. No. Name Description Price
HR3-511 1.88 inch wide Crystal Clear Sealing Tape 1.88 inch roll, x 43.7 yard, $7.00
with cutter
HR4-511 1.88 inch wide Crystal Clear Sealing Tape 1.88 inch x 60 yard roll $9.00
HR4-506 3 inch wide Crystal Clear Sealing Tape 3 inch x 55 yard roll $10.00
f e a t u r e s
Plate 1.88 inch 3 inch
Corning

Cryschem

Cryschem

24-1 SBS
Douglas Instruments
CrystalClear Strips

Greiner
Intelli-Plate

MRC/Swissci

X
X
X
X
X
X
X
C R Y S T A L C L E A R S E A L I N G T A P E
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101
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d e s c r i p t i o n
A 38 m soft non-permeable aluminum foil sealing film with
strong medical-grade adhesive, AlumaSeal II sealing films
eliminate the need for heat-sealing devices or mats during the
resealing of reagents in Deep Well blocks. Each sealing film
measures 82.6 x 142.9 mm and offers sufficient sealing area for
all 96 Deep Well blocks. Length between the perforations with
end tabs removed is 125.4 mm. Compared to other aluminum
foils, AlumaSeal II has less tendency to roll back on itself when
removing the backing paper and conforms well to the plate during application.
AlumaSeal II is a soft, pierceable adhesive film designed for the convenient and rapid sealing of reagent
blocks. A multiple split backing with two end tabs allows for easy, accurate positioning and secure seal-
ing. The use of an adhesive sealing film minimizes evaporation and helps to prevent well-to-well cross
contamination in reagent blocks. AlumaSeal II films are easily pierced by pipettte tips or robotic probes
or piercing tools for direct reagent recovery without significant gumming by adhesive.
Polypropylene mat snap seals MASTERBLOCK 96 Deep Well plate.
n
Sealing film used to reseal HT format
screen kits
n
excellent seal
n
Film conforms to raised chimney wells
n
easily pierceable with single or multichannel
pipetters and robotic probes
n
Less evaporation than clear films
n
Heat & cold resistant, recommended for
temperatures from -80 C to +120 C
n
Certified DNase-, RNase-,
and nucleic-acid-free
n
excellent barrier properties, virtually no
reagent evaporation or drying
n
Snap seal MaSTeRBLOCk 96 Deep Well
plate.
n
Polypropylene
a p p l i c a t i o n
a p p l i c a t i o n
f e a t u r e s
f e a t u r e
Order Information
Cat. No. Name Description Price
HR8-069 AlumaSeal II Sealing Film 100 pack $65.00
HR4-525 Sealing Film Applicator each $5.00
A L U M A S E A L I I S E A L I N G F I L M A N D A P P L I C A T O R
Order information
Cat. No. Name Description Price
HR3-103 Cap Mat for MASTERBLOCK 50 pack $160.00
C A P M A T F O R M A S T E R B L O C K

d e s c r i p t i o n
The Robolid combines the sealing abilities of a silicone rubber mat
and the automation friendliness of a polystyrene lid. Essentially,
it is a silicone mat with tapered well plugs specially bonded to a
polystyrene plate cover. It is compatible with a wide range of liquid
handling automation. The Robolid can be used to cover a 96 Deep
Well block for low evaporation, short-term storage, or to minimize
evaporation or contamination during pipetting. The Robolid is
compatible with the 96 Deep Well blocks used for Hampton Research HT kits such as Crystal Screen
HT, Index HT, and SaltRx HT.
a p p l i c a t i o n
f e a t u r e s
Order Information
Cat. No. Name Description Price
HR3-111 Robolid 25 pack $223.00
R O B O L I D

n
Deep Well block cover for HT screens
n
Lid dimensions are suitable for a wide range
of standard microplates and liquid handling
work stations
n
No well-to-well contamination to ensure
multiple application and removal of lid
n
Silicone sealing plugs are suitable with
organic solvents and low extractables
n
Designed for low evaporation under standard
storage conditions from -20 to 37C
D I A L Y S I S B U T T O N S

, M E M B R A N E S & A P P L I C A T O R S
Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com 102
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In the dialysis method, the sample in question is separated from
the precipitant by a semi-permeable membrane which allows
small reagent molecules to pass but prevents biological macro-
molecules from crossing the membrane.
1-3
Dialysis Buttons are
either machined from transparent perspex or injection molded
from polystyrene, and are the size of a small button. The sample is
placed in this chamber so as to create a slight dome of liquid at the
top of the button. A dialysis membrane is placed over the top of
the button/sample and is held in place with an O-ring. The O-ring
is held in place by a groove in the Dialysis Button. Dialysis Buttons
are supplied with O-rings. The golf tee is supplied with the 5 l to
100 l buttons.
References
1. McPherson, A., Preparation and Analysis of Protein Crystals, Krieger Publishing, 88-91 (1992).
2. Durcruix, A, and Giege, R., Crystallization of Nucleic Acids and Proteins, A Practical Approach,
Oxford University Press, 78-82 (1992).
3. Zeppenzauer, M., et al., Acta Chem Scan (1967) 21, 1009.
n
Crystallization by dialysis, protein
folding, & small volume sample dialysis
n
Low volume dialysis
n
Fits in 24 well plates
n
Use over and over again
a p p l i c a t i o n
f e a t u r e s
Order Information
Cat. No. Name Description Price
HR3-336 Dialysis Buttons Sampler 5 of each size $88.00
HR3-314 5 l Dialysis Button 50 pack $88.00
HR3-316 10 l Dialysis Button 50 pack $88.00
HR3-318 15 l Dialysis Button 50 pack $88.00
HR3-320 20 l Dialysis Button 50 pack $88.00
HR3-322 25 l Dialysis Button 50 pack $88.00
HR3-326 50 l Dialysis Button 50 pack $88.00
HR3-328 100 l Dialysis Button 50 pack $88.00
HR3-330 200 l Dialysis Button 50 pack $88.00
HR3-332 350 l Dialysis Button 50 pack $88.00
How To: Using
Dialysis Buttons
Description
Crystallization by dialysis is an easy variation to the
typical vapor diffusion method used to grow crystals.
In the dialysis method, the sample in question is sep-
arated from the precipitant by a semi-permeable
membrane which allows small molecules such as
ions, additives, buffers, and, salts to pass but pre-
vents biological macromolecules from crossing the
membrane. Equilibration kinetics depend upon the
molecular weight cutoff of the dialysis membrane,
the precipitant, the ratio of the volume, the concen-
tration of the components inside and outside of the
dialysis cell, and the geometry of the cell.
Step 1 - Place sample into dialysis cell
Pipet sample into the dialysis chamber. Amount
of sample can vary depending on the size of the
Dialysis Button.
Step 2 - Placing the membrane
Choose a dialysis membrane that best suits your
molecular weight cutoff of choice. Place the mem-
brane across the top of the Dialysis Button. Using a
golf tee or applicator, press down on the membrane
firmly, making sure no air is trapped between the
membrane and dialysis chamber. Roll the O-ring
over the golf tee and onto the Dialysis Button. Make
sure the O-ring is placed securely in the O-ring sup-
port, holding down the membrane.
Step 3 - Begin dialysis
Place the sealed Dialysis Button into a VDX Plate
or Linbro

Plate. Fill the reservoir with crystalliza-

tion reagent and let the dialysis begin.
Dialysis Chamber
Dialysis Button
O-Ring Support
O-Ring
Golf Tee
Cover Slide (or Sealing Tape)
Well of VDX
Crystallization Plate
Reservoir Solution
Dialysis Button
Vacuum
Grease
D I A L Y S I S B U T T O N S

103
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The Applicator makes easier work of applying the dialysis mem-
brane and O-ring to Dialysis Buttons. It positions and holds the
dialysis membrane on the buttons and also allows the easy applica-
tion and position of the O-ring to secure the dialysis membrane
onto the Dialysis Button. The Applicator for Dialysis Buttons is
manufactured of glass and is available in two sizes. The Small
Applicator (HR4-348) is designed for use with 5 to 100 l Dialysis
Buttons. The Large Applicator (HR4-350) is designed for use with
200 and 350 l Dialysis Buttons.
n
application of dialysis membrane to Dialysis
Buttons
n
Manufactured of glass
n
Two sizes available
n
Small applicator (5 - 100 l Dialysis
Buttons)

n
Large applicator (200 & 350 l Dialysis
Buttons)
a p p l i c a t i o n
f e a t u r e s
Order Information
Cat. No: Name: Description Price
HR4-348 Small Applicator for use with 5 to 100 l Dialysis Buttons $47.00
HR4-350 Large Applicator for use with 200 and 350 l Dialysis Buttons $47.00
A P P L I C A T O R F O R D I A L Y S I S B U T T O N S

d e s c r i p t i o n
Spectra/Por

regenerated cellulose dialysis membrane discs are

33 mm, circular, pre-cut membranes available with the following
molecular weight cutoffs:
3,500 MW cutoff
6,000 - 8,000 MW cutoff
12,000 - 14,000 MW cutoff
n
Membranes for dialysis
a p p l i c a t i o n
D I A L Y S I S M E M B R A N E D I S C S

F O R B U T T O N S
Order Information
Cat. No. Name Description Price
HR3-338 Dialysis Membrane Discs cutoff 3,500 - 50 pack $195.00
HR3-344 Dialysis Membrane Discs cutoff 6,000 to 8,000 - 50 pack $195.00
HR3-346 Dialysis Membrane Discs cutoff 12,000 to 14,000 - 50 pack $195.00
HR4-348 Small Applicator for use with 5 to 100 l Dialysis Buttons $47.00
HR4-350 Large Applicator for use with 200 and 350 l Dialysis Buttons $47.00
Protein crystals.
Kasumi Kobayashi, Taiji Nakae and Hiroyuki Akama.
The Kitasato Institute, Kanagawa, Japan.

d e s c r i p t i o n
The Granada Crystallization Box (GCB) consists of
four elements made of polystyrene:
1. A reservoir to introduce the gel
2. A guide to hold capillaries
3. A cover
4. A holder to maintain the boxes
The GCB has been designed to be used in four different ways:
1. To grow crystals inside gels under diffusion-controlled mass transport (figure 1).
2. To grow crystals inside capillaries with un-gelled precipitating agent by the counter-diffusion
technique (figure 2).
3. To grow crystals inside capillaries with gelled precipitating agent by the counter-diffusion
technique (figure 3).
4. To grow crystals inside capillaries by the batch method (figure 4).
Capillaries and gel are not included with the Granada Crystallization Box.
The capillaries described on this page are designed for use with the Granada Crystallization Box. The
capillaries are extremely thin walled. The internal diameter for these capillaries is of 0.1, 0.2 and 0.3
mm. The respective external diameters are 0.17, 0.33 and 0.40 mm. The borosilicate glass capillaries
are available in four different lengths of 30, 40, 50 or 100 mm. Although the capillaries are designed
for use with the Granada Crystallization Box they can also be used for general free-interface diffusion
crystallization.
References
1. Granada Crystallisation Box: a new device for protein crystallisation by counter-diffusion techniques. Garcia-Ruiz JM, Gonzalez-Ramirez LA, Gavira JA,
Otalora F. Acta Crystallogr D Biol Crystallogr. 2002 Oct;58 (Pt 10 Pt 1):1638-42.
2. Counterdiffusion methods for macromolecular crystallization. Garcia-Ruiz JM. Methods Enzymol. 2003;368:130-54.
3. A simplified counter diffusion method combined with a 1D simulation program for optimizing crystallization conditions. H. Tanaka, K. Inaka, S. Sugiyama, S.
Takahashi, S. Sano, M. Sato and S. Yoshitomi. J. Synchrotron Rad. (2004). 11, 45-48.
n
Crystallization using gels in capillaries
n
Crystallization inside glass or quartz
capillaries
n
Can be used with or without gel
n
Counter-diffusion or batch method
n
Optically clear polystyrene
a p p l i c a t i o n
Order Information
Cat. No. Name Description Price
HR3-194 Granada Crystallization Box 20 plates and 1 holder $261.00
HR4-677 Round Capillary 100 mm x 0.1 mm - 25 pack $30.00
HR4-678 Round Capillary 100 mm x 0.2 mm - 25 pack $30.00
HR4-679 Round Capillary 100 mm x 0.3 mm - 25 pack $30.00
HR4-680 Round Capillary 50 mm x 0.1 mm - 25 pack $26.00
HR4-681 Round Capillary 50 mm x 0.2 mm - 25 pack $26.00
HR4-682 Round Capillary 50 mm x 0.3 mm - 25 pack $26.00
HR4-683 Round Capillary 40 mm x 0.1 mm - 25 pack $26.00
HR4-684 Round Capillary 40 mm x 0.2 mm - 25 pack $26.00
HR4-685 Round Capillary 40 mm x 0.3 mm - 25 pack $26.00
HR4-686 Round Capillary 30 mm x 0.1 mm - 25 pack $26.00
HR4-687 Round Capillary 30 mm x 0.2 mm - 25 pack $26.00
HR4-688 Round Capillary 30 mm x 0.3 mm - 25 pack $26.00
HR8-092 LM Agarose 10 g bottle $58.00
figure 1
figure 2
figure 3
figure 4
f e a t u r e s
m a t e r i a l
G R A N A D A C R Y S T A L L I Z A T I O N B O X

& C A P I L L A R I E S
104
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Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com
Crystals of the Deinococcus radiodurans 50S ribosome subunit.
From the group of Paola Fucini, Max Planck Institute
for Molecular Genetics, Berlin, Germany.
Protein crystals.
Kasumi Kobayashi, Taiji Nakae and Hiroyuki Akama.
The Kitasato Institute, Kanagawa, Japan.
Deoxyribonuclease crystals grown in dAMP, GDP,
naladixic acid, 15% w/v Polyethylene glycol 3,350.
John Day, University of California Irvine, USA.
t o o l s , s e e d i n g & r e s i n
Crystal image UFO.
Chongping Chen, Dr. Jaffe Lab, Fox Chase Cancer Research Center.
T A B L E O F C O N T E N T S
108 m i c r o - t o o l s

109 m i c r o - t o o l s I I

110 - 113 f o r c e p s
113 c r y s t a l p r o b e

113 c r y s t a l p e n c i l

114 s e e d b e a d

115 s e e d i n g t o o l
115 c h e l a t i n g r e s i n
P A G E S
n
Precision instruments for crystal manipulation
n
Made from chemical resistant,
hardened tool steel
n
Interchangeable tools tips
M I C R O - T O O L S

Crystal shown being cut using the Hampton Research

Micro-Knife.
Courtesy of Lisa Edberg.
University of Alabama, Birmingham
Center for Macromolecular Crystallography
Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com
a p p l i c a t i o n
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108
s u c c e s s s t o r y
Micro-Tools are the smallest available precision instruments for
laboratory use. These tools are realistically proportioned for mac-
romolecular and small molecule crystallization work. The high
quality, precision Micro-Tools ease microscopic manipulations.
The eight specifically engineered Micro-Tools are useful in a vari-
ety of applications in the crystallization lab. Eight tools, a handle
and wood storage box.
Micro-Tools are designed to be versatile as well as useful for
specific crystallization manipulations. For example, the Micro-Scale is very useful for measuring
crystal dimensions as well as serving as a way to record the dimensions by photography for reference
or publication. The Micro-Chisel and Micro-Spade are helpful for separating and splitting blade and
whisker clusters during seeding or mounting. The Micro-Prober and Micro-Needle come in handy
when probing precipitate or when manipulating small or large crystals for seeding as well as during
mounting. The Micro-Spatula can be used to move crystals as well as split single crystals or clusters.
The Micro-Scraper is helpful when working with very small crystals.
The Micro-Tools Set is indispensable during crystal observation, manipulation, seeding, and mount-
ing. The set includes a single anodized aluminum handle which holds each of the eight unique tips,
all supplied in an attractive alderwood instrument case. The case provides protective storage of the
Micro-Tools as well as a convenient carrying case.
The tips are constructed from hardened tungsten steel and are resistant to most common reagents
used for crystal growth. The threaded tool tips are easily interchanged with the threaded aluminum
handle. The diameter of the tool shaft is 0.250 mm (0.010 inches). The actual dimension at the very
tip of the tool will vary with the particular tool design.
Order Information
Cat. No. Name Description Price
HR4-811 Micro-Tools Set each $298.00
HR4-813 Micro-Scale each $55.00
HR4-815 Micro-Spade each $48.00
HR4-817 Micro-Prober 45 each $48.00
HR4-819 Micro-Scraper each $54.00
HR4-821 Micro-Chisel each $54.00
HR4-823 Micro-Knife 20 each $54.00
HR4-825 Micro-Spatula each $52.00
HR4-827 Micro-Needle each $46.00
HR4-829 Micro-Tool Handle each $33.00
HR4-831 Micro-Tools Carrying Case each $56.00
1
2
3
4
5
1.
2.
3.
7.
4.
6.
8.
5.
Micro-Tool Set:
1. Micro-Scraper
2. Micro-Spatula
3. Micro-Needle
4. Micro-Scale
5. Micro-Spade
6. Micro-Knife 20
7. Micro-Chisel
8. Micro-Prober 45
109
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Precision instruments for crystal manipulation
n
Made from chemical resistant, tungsten steel
and hardened tool steel
n
Interchangeable tools tips
n
8 unique tools designed to extend the
original Micro-Tools set
a p p l i c a t i o n
f e a t u r e s
d e s c r i p t i o n
Micro-Tools II Set is an extension to the smallest available preci-
sion instruments for laboratory use. These tools are realistically
proportioned for macromolecular and small molecule crystal-
lization work. The high quality, precision Micro-Tools II ease
microscopic manipulations. The eight specifically engineered
Micro-Tools II are useful in a variety of applications in the crystal-
lization lab. Eight tools, a handle and wood storage box.
The Micro-Tools II are designed to be versatile as well as useful
for specific crystallization manipulations. For example, the Micro-
Manipulator is handy for manipulating long, thin blades or for sep-
arating long, thin blade clusters. Awkward positions encountered
during crystal manipulation can sometimes be avoided using tools
with an unusual and flexible approach angle such as the Micro-
Prober 90 and the Micro-Hook 90. Seeding experiments can be performed using the Micro-Brush.
The Ultra Micro-Needles are extremely delicate tools crafted from 5 micron radius tungsten that are
very handy for seeding as well as manipulations requiring a soft, fine, and controlled touch.
The Micro-Tools II Set is indispensable during crystal observation, manipulation, seeding, and mount-
ing. The set includes a single anodized aluminum handle which holds each of the eight unique tips,
all supplied in an attractive alderwood instrument case. The case provides protective storage of the
Micro-Tools as well as a convenient carrying case.
The tips are constructed from hardened tungsten steel and are resistant to most common reagents
used for crystal growth. The threaded tool tips are easily interchanged with the threaded aluminum
handle. The diameter of the tool shaft is 0.250 mm (0.010 inches). The actual dimension at the very
tip of the tool will vary with the particular tool design.
Order Information
Cat. No. Name Description Price
HR4-837 Micro-Tools II Set each $298.00
HR4-839 Micro-Manipulator each $49.00
HR4-841 Micro-Knife 45 each $49.00
HR4-843 Micro-Prober 90 each $48.00
HR4-845 Micro-Brush each $44.00
HR4-847 Micro-Hook 90 each $49.00
HR4-849 Ultra Micro-Needle, Straight each $41.00
HR4-851 Ultra Micro-Needle 30 each $49.00
HR4-853 Ultra Micro-Needle 90 each $49.00
HR4-829 Micro-Tool Handle each $33.00
HR4-831 Micro-Tools Carrying Case each $56.00
M I C R O - T O O L S I I

HR4-821
1.
2.
3.
4.
5.
6.
7.
8.
Micro-Tool II Set :
1. Manipulator
2. Micro-Brush
3. Micro-Knife 45
4. Micro-Prober 90
5. Ultra Micro-Needle, straight, tungsten, 5 micron radius
6. Ultra Micro-Needle, bent 30, tungsten, 5 micron radius
7. Micro-Hook 90
8. Ultra Micro-Needle, bent 90, tungsten, 5 micron radius
n
Length - 115 mm
n
Tip - 9.5 mm to point
n
Length - 150 mm
n
Tip - 10.3 mm to point
n
Length - 137 mm
n
Tip - 2.5 mm to point
n
Length - 119 mm
n
Head Length - 11.5 mm
n
Head Width - 6.1 mm
U L T R A - F I N E F O R C E P
S L I D E - T E N S I O N F O R C E P
4 5 A N G L E D F O R C E P
3 0 T I P A N G L E D F O R C E P
F O R C E P S
Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com
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110
f e a t u r e s
f e a t u r e s
f e a t u r e s
f e a t u r e s
High quality, precision straight forceps with an extra fine
point. Smooth, non-serrated tip and handle. Great for
mounting CryoLoops.
Straight forcep with adjustable "locking" tension bar.
Increase the width of the tip opening by setting the bar to
the desired position. Slide bar can be positioned to "lock"
forceps and hold cover slides, CryoLoops, MicroTubes, and
other small parts.
All purpose angled forcep with blunt end. Tip has serration
for firm grip. Good for retrieving. 45 angle makes for easy
manipulation. Especially nice for flipping cover slides. Has
serrated non-slip handle.
Smooth, wide, angled, flat tip that will leave delicate items
undamaged, unlike tweezers with sharp points. Tip is
angled down 30 from the handle. Has serrated, non-slip
handle. Good for handling cover slides and filter paper.
d e s c r i p t i o n
d e s c r i p t i o n
d e s c r i p t i o n
d e s c r i p t i o n
Order Information
Cat. No. Name Description Price
HR4-855 Ultra-Fine Forcep each $10.00
Order Information
Cat. No. Name Description Price
HR4-857 Slide-Tension Forcep each $8.00
Order Information
Cat. No. Name Description Price
HR4-859 45 Angled Forcep each $10.00
Order Information
Cat. No. Name Description Price
HR4-861 30 Tip Angled Forcep each $8.00
111
n
Length - 140 mm
n
Tip Width - 2.5 mm
n
Length - 200 mm
n
Tip Width - 2.5 mm
n
Length - 300 mm
n
Tip Width - 2.5 mm
S M A L L F O R C E P
M E D I U M F O R C E P
E X T R A L A R G E F O R C E P
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f e a t u r e s
f e a t u r e s
All purpose straight forcep with blunt end. Tip has serrated
surface for firm grip. Good for retrieving. Has serrated non-
slip handle.
All purpose straight forcep with blunt end. Tip has serrated
surface for firm grip. Good for retrieving. Has serrated non-
slip handle.
All purpose straight forcep with blunt end. Tip has serrated
surface for firm grip. Good for retrieving. Has serrated non-
slip handle.
d e s c r i p t i o n
d e s c r i p t i o n
d e s c r i p t i o n
Order Information
Cat. No. Name Description Price
HR4-863 Small Forcep each $9.00
Order Information
Cat. No. Name Description Price
HR4-865 Medium Forcep each $13.00
Order Information
Cat. No. Name Description Price
HR4-869 Extra Large Forcep each $10.00
F O R C E P S
Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com
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n
Length - 113 mm
n
Tip Length - 31 mm
n
Tip Width - 3.1 mm to point
n
Length - 113 mm
n
Tip Length - 10.9 mm
n
Tip Width - 3.5 mm to point
n
Length - 113 mm
n
Tip Length - Curved to point
n
Length - 156 mm
n
Tip Length - 18 mm
n
Tip Width - 1.7 mm
S T R A I G H T M I C R O F O R C E P
4 5 T I P A N G L E D F O R C E P
C U R V E D T I P M I C R O F O R C E P
A N G L E D T I P L O C K I N G F O R C E P
f e a t u r e s
f e a t u r e s
f e a t u r e s
f e a t u r e s
Ultra-thin, smooth, pointed tip for very precise work. Great
for retrieving or holding small parts. Smooth, non-serrated
handle and tip.
Straight forceps with angled tip. Easy to use forceps when
space is limited. Tip is slanted at a 45 angle. Smooth, non-
serrated handle and tip.
Ultra-thin, smooth, pointed tip for very precise work. Great
for retrieving or holding small parts. Smooth, non-serrated
handle and tip.
All purpose angled tip forcep. Tip has serrated surface for
firm grip. Good for retrieving. When closed and squeezed,
forcep locks to clamp and hold item in place. Lock is
released by moving lock toward tip with thumb. Serrated,
non-slip handle. Angled for easy manipulation.
d e s c r i p t i o n
d e s c r i p t i o n
d e s c r i p t i o n
d e s c r i p t i o n
Order Information
Cat. No. Name Description Price
HR4-871 Straight Microforcep each $13.00
Order Information
Cat. No. Name Description Price
HR4-879 Curved Tip Microforcep each $9.00
Order Information
Cat. No. Name Description Price
HR4-881 Angled Tip Locking Forcep each $13.00
Order Information
Cat. No. Name Description Price
HR4-875 45 Tip Angled Forcep each $8.00
113
a p p l i c a t i o n s
a p p l i c a t i o n s
n
Crystal manipulation
n
Seeding
n
Stainless steel probe with plastic handle
n
Crystal manipulation
n
Seeding
n
Crystal mounting
f e a t u r e s
Disposable Crystal Probe manipulators made from a 0.12
mm x 30 mm stainless steel, pointed-end probe attached
to a 20 mm plastic handle. Useful for breaking apart blade
clusters and general crystal manipulation. Small size and
disposable format makes them convenient for trips to the
synchrotron.
Useful for manipulating crystals for seeding and cryocrystal-
lography. The 0.7 mm tip accepts the Hampton Research
Mounted CryoLoops or MicroTubes with your own
cryoloop which creates an easy to hold, comfortable,
and convenient tool for manipulating crystals. CryoLoops,
MicroTubes, and Mounted CryoLoops sold separately.
d e s c r i p t i o n
d e s c r i p t i o n
C R Y S T A L P E N C I L

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C R Y S T A L P R O B E

n
Length - 156 mm
n
Tip Width- 1.7 mm
L O C K I N G F O R C E P
f e a t u r e s
All purpose straight forcep. Tip has serrated surface for firm
grip. Good for retrieving. When closed and squeezed, for-
cep locks to clamp and hold item in place. Lock is released
by moving lock toward tip with thumb. Serrated, non-slip
handle.
d e s c r i p t i o n
Order Information
Cat. No. Name Description Price
HR4-883 Locking Forcep each $13.00
Order Information
Cat. No. Name Description Price
HR4-217 Crystal Probe 12 pack $15.00
Order Information
Cat. No. Name Description Price
HR4-835 Mechanical Crystal Pencil each $3.00
a p p l i c a t i o n
n
Generate seeds of protein crystals
n
Easily generate consistent seed stocks
n
Use serial dilution to control the number
of seeds introduced into the drop
f e a t u r e s
S E E D B E A D

Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com
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The Seed Bead kit is used to create a seed stock for
performing subsequent seeding experiments. The Seed
Bead kit contains 24 Seed Beads manufactured from PTFE,
individually contained in a special 1.5 ml microcentrifuge
tube.
Seeding allows one to grow crystals in the metastable zone.
Crystallization in this zone provides control, reproducibility
and an improved likelihood of a successful crystallization experiment. Also, crystals can grow from seeds
but cannot spontaneously nucleate. By placing a seed or solution of seeds in a drop which is saturated
to the metastable zone, one can use the seeds to grow larger single crystals. By controlling the number
of seeds introduced into the drop, one can control the number of crystals grown. It is not practically
possible to measure and know the number of seeds introduced to a drop, but by performing serial dilu-
tions from a concentrated seed stock, one can control the number of crystals grown in the drop.
Using the Seed Bead kit, one can create crystal seed stock for subsequent seeding experiments. Crystals
are placed in the microcentrifuge tube with the PTFE Seed Bead and either vortexed or sonicated to
generate a seed stock. Then by performing serial dilutions, one can control the number and size of
crystals in the experiment.
The Seed Bead kit is useful for the preparation of seed stocks for automated and semi-automated
microseeding (D'Arcy 2007, Harlos 2008).
Each kit contains 24 special microcentrifuge tubes with Seed Beads. Crystallization accessories are sold
separately.
References
1. Stura, E.A., Wilson, I.A., Methods: A Companion to Methods in Enzymology (1990) 1, 38-49.
2. Stura, E.A., Wilson, I.A., "Seeding Techniques" in Crystallization of Nucleic Acids and Proteins: A Practical Approach. Oxford University Press (1992) 99-126.
3. Luft, J.R., DeTitta, G.T., Poster: Using Ultrasound in the preparation of Micro-Seed Stock for the Crystallization of Macromolecules, 1997 ACA Meeting, St
Louis, MO. Publication in preparation.
4. J.R. Luft and G.T. DeTitta, Methods in Enzymology (1997) 276, 110-131.
5. Structure of an orthorhombic form of xylanase II from Trichoderma reesei and analysis of thermal displacement. Watanabe et al. Acta Cryst. (2006). D62,
784-792.
6. Crystallization and preliminary crystallographic analysis of p40phox, a regulatory subunit of NADPH oxidase. K. Honbou, S. Yuzawa, N. N. Suzuki, Y.
Fujioka, H. Sumimoto and F. Inagaki. Acta Cryst. (2006). F62, 1021-1023 (Used seed bead to optimize)
7. Purification, crystallization and preliminary X-ray diffraction study of human ribosomal protein L10 core domain. Yuji Kobayashi et al. Acta Cryst. (2007). F63,
950952
8. Semi-automated microseeding of nanolitre. Thomas S. Walter, Erika J. Mancini, Jan Kadlec, Stephen C. Graham, Rene Assenberg, Jingshan Ren, Sarah
Sainsbury, Raymond J. Owens, David I. Stuart, Jonathan M. Grimes and Karl Harlos Acta Cryst. (2008). F64, 1418
9. An automated microseed matrix-screening method. Allan DArcy,a* Frederic Villard and May Marsh. Acta Cryst. (2007). D63, 550554
d e s c r i p t i o n
Order Information
Cat. No. Name Description Price
HR2-320 Seed Bead kit 24 tubes with Seed Beads $56.00
Simply pipet your sample into the special 1.5 ml microcentrifuge
tube containing the Seed Bead. Vortex the sample and youve
got seed stock.
115
a p p l i c a t i o n
a p p l i c a t i o n
n
Streak Seeding
n
Natural fiber
n
Built-in handle/cover
n
Remove trace metals from water, reagents,
& samples
n
Scavenge multivalent metal ion contaminants
n
Remove nickel from sample following
purification over a nickel column
f e a t u r e s
f e a t u r e s
During streak seeding, one touches the Seeding Tool to
crystalline material to dislodge, remove and transfer small
crystals (seeds) to a drop that will support the growth of
potentially larger and more perfect crystals. A seed can pro-
vide a template on which additional macromolecules can
assemble and under the proper conditions, grow to form
a large single crystal. Using seeding can avoid problems
associated with growing crystals from spontaneous nucleation. Seeds can grow into larger crystals in
the metastable region of the solubility curve, which is a region of lower, relative supersaturation. One
can also streak seed from phase separation or amorphous material as a diagnostic to confirm whether
the material is crystalline. The Seeding Tool is a 1 cm natural fiber attached to a stainless steel pin and
plastic handle. The Seeding Tool is supplied with a cover to protect the fiber and when the cover is
placed on the back side of the Seeding Tool, it makes an excellent handle.
References
1. Seeds to crystals. Terese Bergfors. Journal of Structrual Biology 142 (2003) 66-76.
2. Applications of the streak seeding technique in protein crystallization. Stura and Wilson. Journal of CrystaL Growth, 110 (1991) 270-282.
Chelating Resin can be used to remove trace metals from water,
reagents, and macromolecular samples
1-3
.
Chelating Resin has a high preference for mercury, copper, nickel,
lead, zinc, cobalt, cadmium, iron, manganese, barium, calcium,
strontium, and magnesium. Its selectivity for divalent ions over
monovalent ions is high (5,000:1) and it has a very high attraction
for transition metals, even in concentrated salt solutions. The resin
will scavenge multivalent metal ion contaminants without altering the concentration of nonmetallic
ions. In most cases the resin does not have any effect on protein concentration or activity.
Chelating Resin is a high purity, 100-200 mesh resin. This resin is a styrene divinylbenzene copolymer
containing paired iminodiacetate ions which act as chelating groups in binding polyvalent metal ions.
The nominal capacity is 2.0 meq/dry gram or 0.4 meq/ml resin bed. The density is 0.65 g/ml. The resin
can be classed with weakly acidic cation exchange resins due to the presence of carboxylic acid groups
but differs from ordinary exchange resins due to its high selectivity for metal ions and high bond
strength. The resin is stable over the entire pH range and functionally active from pH 2 to 14. The resin
is stable for at least two years when stored in the original sealed container at 23C.
References
1. Removal of Nickel from protein solutions following purification over a Nickel column. Hassell, A., Recent Advances in Macromolecular Crystallization,
Bischenberg, FRANCE (1997).
2. Dunn, M.F., Pattison, S.E., Storm, M.C., and Quiel, E., Removal of metals from enzyme solutions., Biochem. (1980) 19, 718.
3. Ray, W.J., Burgner, J.W., and Post, C.B., Purification of NMR reagents., Biochem. (1990) 29, 2770.
d e s c r i p t i o n
d e s c r i p t i o n
Order Information
Cat. No. Name Description Price
HR8-133 Seeding Tool 5 pack $30.00
Order Information
Cat. No. Name Description Price
HR2-312 Chelating Resin 10 g bottle $38.00
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S E E D I N G T O O L
C H E L A T I N G R E S I N

Here is a recipe to try:

Mosaicity is about 0.5
t0 0.6
Reagent:
Crystal Screen Cryo
Reagent 23
Mix equal amounts of
Glucose Isomerase
and reagent. Vapor
diffusion method.

Mount crystal in
CryoLoop.

Mosaicity may be
a bit more in this
reagent and the
unit cell will shrink a
C r y s t a l l i z a t i o n S c r e e n s c r y o c r y s t a l l o g r a p h y
A star of H. influenzae carbonic anhydrase crystals.
Roger S. Rowlett, Department of Chemistry, Colgate University, Hamilton, New York, USA.
T A B L E O F C O N T E N T S
118 - 120 c r y s t a l c a p

h t s y s t e m s
121 - 122 c r y s t a l c a p

m a g n e t i c s y s t e m s
122- 123 c r y s t a l c a p

s y s t e m s
124 - 127 c r y o l o o p s & m i c r o t u b e s
128 - 132 c r y o t o o l s
133 c a n e s , s l e e v e s & c o d e r s
134 - 135 d e w a r s
P A G E S
Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com 118
n
Cryocrystallography
n
Specially designed vial with magnetic ring

n
Cap magnetically attaches to vial
n
Copper jacket to reduce icing on MicroTube
n
Vented vial sold separately
n
No threads
n
Bar coded, color coded, alphanumeric,
magnetic cap
n
Assembled with Mounted CryoLoops,
ready-to-use; vials sold separately
n
Cap features a flat ledge used by grippers
such as SSRL Stanford Automated Mounting
System (SAM)
a p p l i c a t i o n
f e a t u r e s
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C R Y S T A L C A P

H T

S Y S T E M S
CrystalCap Copper Magnetic HT is designed to prevent ice forma-
tion along the copper pin/MicroTube in a cryostream where
the flow is positioned perpendicular, at an angle, or non-collinear
to the MicroTube. The CrystalCap Copper Magnetic HT is a non-
threaded, 1.8 ml (approximate) cryo vial storage container with a
specially engineered cap/pin for cryocrystallography. Each cap is
manufactured from an alloy base which allows the cap to be mag-
netically secured to a magnetic base in the goniometer head. Into
the alloy cap is threaded and bonded a solid, 3 mm diameter copper pin. The end of the copper pin
is machined to an aerodynamic 60 and is assembled with a MicroTube and CryoLoop. A ring magnet
is molded into the top end of the vial so that when the cap is positioned in the vial, the ring magnet
holds the cap on the vial during cryogenic storage. The vial has two holes for venting. The 18 mm
designates the distance between the top of the copper pin and the bottom of the CrystalCap Copper
Magnetic where contact is made with the magnetic base.
For crystal transfer under cryo temperature use the 18 mm CryoTong (catalog number HR4-637 for
the 110 mm overall tool length or catalog number HR5-112 for the 180 mm overall tool length).
The cap features a two dimensional bar coded and alphanumeric 16 x 16 data matrix. Each cap is also
color coded.
HR8-181 CrystalCap Copper Magnetic HT is a two dimensional bar coded and alphanumerically
labeled cap with copper pin; no Mounted CryoLoop and no vial.
Note: Caps with Mounted CryoLoops are sold without vials. Vials sold separately.
Color Coded Cap CryoLoop Size
Red 0.025 - 0.05 mm
Green 0.05 - 0.1 mm
Yellow 0.1 - 0.2 mm
Blue 0.2 - 0.3 mm
Blue/Red 0.3 - 0.4 mm
Green/Red 0.4 - 0.5 mm
Yellow/Red 0.5 - 0.7 mm
Yellow/Green 0.7 - 1.0 mm
The CrystalCap Copper Magnetic HT is not compatible with the EMBL/ESRF SC3 sample changer. For
EMBL/ESRF SC3 and other SPINE sample changers please use the CrystalCap HT.
d e s c r i p t i o n
Order Information
Cat. No. Name Description Price
HR8-173 CrystalCap Copper Magnetic HT 0.025 - 0.05 mm CryoLoop - 30 pack $195.00
HR8-174 CrystalCap Copper Magnetic HT 0.05 - 0.1 mm CryoLoop - 30 pack $195.00
HR8-175 CrystalCap Copper Magnetic HT 0.1 - 0.2 mm CryoLoop - 30 pack $195.00
HR8-176 CrystalCap Copper Magnetic HT 0.2 - 0.3 mm CryoLoop - 30 pack $195.00
HR8-177 CrystalCap Copper Magnetic HT 0.3 - 0.4 mm CryoLoop - 30 pack $195.00
HR8-178 CrystalCap Copper Magnetic HT 0.4 - 0.5 mm CryoLoop - 30 pack $195.00
HR8-179 CrystalCap Copper Magnetic HT 0.5 - 0.7 mm CryoLoop - 30 pack $195.00
HR8-180 CrystalCap Copper Magnetic HT 0.7 - 1.0 mm CryoLoop - 30 pack $195.00
HR4-904 CrystalCap Magnetic Vial Vial ONLY - 30 pack $84.00
HR8-181 CrystalCap Copper Magnetic HT Cap ONLY - 30 pack $130.00
C R Y S T A L C A P

C O P P E R M A G N E T I C H T
2
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2
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m
m
0.65 mm
9.7 mm
12 mm
9.8 mm
1.5 mm
CrystalCap Copper Magnetic HT
119
n
Cryocrystallography
n
Specially designed vial with magnetic ring

n
Cap magnetically attaches to vial
n
Copper jacket to reduce icing on MicroTube
n
Vented vial sold separately
n
No threads
n
Bar coded, color coded, alphanumeric,
magnetic cap
n
Assembled with Mounted CryoLoops,
ready-to-use; vials sold separately
n
Compatible with ALS style sample handlers,
mounters
n
Cap has ledge free, conical shape used by
grippers in ALS - style automounters. For
use at LBNL Berkeley Center for Structural
Biology, with the PXRR automounter at
Brookhaven NSLS, and at CHESS
a p p l i c a t i o n
f e a t u r e s
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CrystalCap Copper Magnetic ALS HT is designed to prevent ice
formation along the copper pin/MicroTube in a cryostream
where the flow is positioned perpendicular, at an angle, or non-
collinear to the MicroTube. The CrystalCap Copper Magnetic
ALS HT is a non-threaded, 1.8 ml (approximate) cryo vial storage
container with a specially engineered ALS compatible cap/pin for
cryocrystallography. Each ALS style cap is manufactured from an
alloy base which allows the cap to be magnetically secured to a
magnetic base in the goniometer head. Into the alloy cap is threaded and bonded a solid, 3 mm
diameter copper pin. The end of the copper pin is machined to an aerodynamic 60 and is assembled
with a MicroTube and CryoLoop or is available without a MicroTube and CryoLoop (HR8-182). A ring
magnet is molded into the top end of the vial so that when the cap is positioned in the vial, the ring
magnet holds the cap on the vial during cryogenic storage. The vial has two holes for venting. The
18 mm designates the distance between the top of the copper pin and the bottom of the CrystalCap
Copper Magnetic where contact is made with the magnetic base.
For crystal transfer under cryo temperature use the 18 mm CryoTong (catalog number HR4-637 for
the 110 mm overall tool length or catalog number HR5-112 for the 180 mm overall tool length).
The cap features a two dimensional bar coded and alphanumeric 16 x 16 data matrix. Each cap is also
color coded.
HR8-182 is a CrystalCap Copper Magnetic ALS HT cap only. No MicroTube, no CryoLoop, no vial.
Note: Caps with Mounted CryoLoops are sold without vials. Vials available separately.
Color Coded Cap CryoLoop Size
Red 0.025 - 0.05 mm
Green 0.05 - 0.1 mm
Yellow 0.1 - 0.2 mm
Blue 0.2 - 0.3 mm
Blue/Red 0.3 - 0.4 mm
Green/Red 0.4 - 0.5 mm
Yellow/Red 0.5 - 0.7 mm
Yellow/Green 0.7 - 1.0 mm
The ALS style automated sample changer accepts the Hampton Research CrystalCap Copper Magnetic
ALS HT.
The CrystalCap Copper Magnetic ALS HT is not compatible with the EMBL/ESRF SC3 sample changer.
For EMBL/ESRF SC3 and other SPINE sample changers please use the CrystalCap HT.
d e s c r i p t i o n
Order Information
Cat. No. Name Description Price
HR8-184 CrystalCap Copper Magnetic ALS HT 0.025 - 0.05 mm CryoLoop - 30 pack $195.00
HR8-186 CrystalCap Copper Magnetic ALS HT 0.05 - 0.1 mm CryoLoop - 30 pack $195.00
HR8-188 CrystalCap Copper Magnetic ALS HT 0.1 - 0.2 mm CryoLoop - 30 pack $195.00
HR8-190 CrystalCap Copper Magnetic ALS HT 0.2 - 0.3 mm CryoLoop - 30 pack $195.00
HR8-192 CrystalCap Copper Magnetic ALS HT 0.3 - 0.4 mm CryoLoop - 30 pack $195.00
HR8-194 CrystalCap Copper Magnetic ALS HT 0.4 - 0.5 mm CryoLoop - 30 pack $195.00
HR8-196 CrystalCap Copper Magnetic ALS HT 0.5 - 0.7 mm CryoLoop - 30 pack $195.00
HR8-198 CrystalCap Copper Magnetic ALS HT 0.7 - 1.0 mm CryoLoop - 30 pack $195.00
HR4-904 CrystalCap Magnetic Vial Vial ONLY - 30 pack $84.00
HR8-182 CrystalCap Copper Magnetic ALS HT Cap ONLY - 30 pack $130.00
C R Y S T A L C A P

C O P P E R M A G N E T I C A L S H T
2
1
.
5

m
m
2
0

m
m
0.65 mm
9.7 mm
12 mm
9.65 mm
1.5 mm
CrystalCap Copper Magnetic ALS HT
C R Y S T A L C A P

H T

S Y S T E M S
Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com 120 Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com
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Cryocrystallography
n
Specially designed vial with magnetic ring

n
Cap magnetically attaches to vial
n
Vented vial sold separately
n
No threads
n
Bar coded, color coded, alphanumeric,
magnetic cap
n
Assembled with Mounted CryoLoops,
ready-to-use; vials sold separately
n
Compatible with automated sample handlers
- SPINE sample changer
- ALS style sample mounters
- Brookhaven National Lab PXRR
automounter
- ESRF automated sample changer
a p p l i c a t i o n
f e a t u r e s
The CrystalCap HT is a complete crystal mount for
manual and automated cryocrystallography. The cap sup-
ports the MicroTube and CryoLoop and is made from a
corrosion resistant magnetic alloy. Chamfered edges on
the cap avoid potential blocking during transfers. The cap
design minimizes material to reduce the cooling/melt-
ing/drying temperature cycle when the sample holder
is transferred. The cap features a two dimensional, bar
coded and alphanumeric 16 x 16 data matrix. The alloy
cap, alloy MicroTube, and synthetic CryoLoop feature an overall sample holder length of 22 mm
(measured from the base of the cap to beam position). The magnetic vial, which is sold separately, is
vented, features chamfered edges for enhanced cap positioning and a magnetic alloy bottom for sta-
bility. The CrystalCap HT is compatible with numerous commercial and academic automated sample
handling systems. It is available as individual components (cap only, vial only, or cap and vial) or as
an assembled crystal mount system (cap, MicroTube, and CryoLoop). The CryoLoop is a 20 micron
diameter, synthetic material.
For crystal transfer under cryo temperature use the 18 mm CryoTong (catalog number HR4-637
for the 110 mm overall tool length or catalog number HR5-112 for the 180 mm overall tool length).
HR8-094 is the Crystal Cap HT cap only, no color code, no two dimensional bar code, no white
background on bottom of cap, no alphanumeric side labeling, and is the cap only, without Mounted
CryoLoop.
Note: Caps with Mounted CryoLoops are sold without vials. Vials available separately.
Color Coded Cap CryoLoop Size
Red 0.025 - 0.05 mm
Green 0.05 - 0.1 mm
Yellow 0.1 - 0.2 mm
Blue 0.2 - 0.3 mm
Blue/Red 0.3 - 0.4 mm
Green/Red 0.4 - 0.5 mm
Yellow/Red 0.5 - 0.7 mm
Yellow/Green 0.7 - 1.0 mm
The CrystalCap HT is compatible with the SPINE sample changer, ALS style sample mounters, the
Brookhaven National Lab PXRR automounter, and the ESRF automated sample changer.
C R Y S T A L C A P

H T ( S P I N E )
d e s c r i p t i o n
Order Information
Cat. No. Name Description Price
HR8-118 CrystalCap HT 0.05 mm CryoLoop - 30 pack $195.00
HR8-120 CrystalCap HT 0.05 - 0.1 mm CryoLoop - 30 pack $195.00
HR8-122 CrystalCap HT 0.1 - 0.2 mm CryoLoop - 30 pack $195.00
HR8-124 CrystalCap HT 0.2 - 0.3 mm CryoLoop - 30 pack $195.00
HR8-126 CrystalCap HT 0.3 - 0.4 mm CryoLoop - 30 pack $195.00
HR8-128 CrystalCap HT 0.4 - 0.5 mm CryoLoop - 30 pack $195.00
HR8-130 CrystalCap HT 0.5 - 0.7mm CryoLoop - 30 pack $195.00
HR8-132 CrystalCap HT 0.7 - 1.0 mm CryoLoop - 30 pack $195.00
HR4-637 CryoTong for (same as CryoTong for 18 mm) - 1 each $55.00
CrystalCap HT
HR5-112 CryoTong for (same as Long CryoTong for 18 mm) - 1 each $61.00
CrystalCap HT
HR8-112 CrystalCap HT Cap ONLY - 60 pack $81.00
HR8-114 CrystalCap HT Vial ONLY - 30 pack $84.00
HR8-116 CrystalCap HT Cap/Vial ONLY - 30 pack $155.00
HR8-094 CrystalCap HT Cap ONLY, without bar coding - 30 pack $77.00
0.65 mm
2
1
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5

m
m
12 mm
CrystalCap HT
C R Y S T A L C A P

M A G N E T I C S Y S T E M S
121
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Cryocrystallography
n
Magnetic cap

n
Specially designed vial with magnetic ring
n
Cap magnetically attaches to vial
n
Vented vial
n
No threads
n
Vial without magnetic base
a p p l i c a t i o n
f e a t u r e s
The CrystalCap Magnetic is a non-threaded, 1.8 ml (approximate)
cryo vial storage container with a specially engineered cap for
cryocrystallography. Each cap is manufactured from an alloy base
which allows the cap to be magnetically secured to a magnetic base
in the goniometer head. A ring magnet is molded into the top end
of the vial so that when the cap is positioned in the vial, the ring
magnet holds the cap on the vial during cryogenic storage. The tip
of the metal cap is ready to accept a MicroTube and CryoLoop
or Mounted CryoLoop. CrystalCap Magnetic is available with or without a vial. The vial contains two
small holes for venting.
CrystalCap Magnetic ALS (HR4-779) is compatible with ALS style sample changers.
In the comparison image above, the CrystalCap Magnetic (notice ledge at bottom of taper) appears on
the left while the CrystalCap Magnetic ALS (no ledge at bottom of taper) appears on the right. Both
cap styles can accept and use the same CrystalCap Magnetic Vial.
Brookhaven National Lab PXRR automounter is compatible with the CrystalCap Magnetic ALS.
The automounter at the Berkeley Center for Structural Biology at Lawrence Berkeley National Labs is
compatible with the CrystalCap Magnetic ALS.
ALS style sample mounters accept the CrystalCap Magnetic ALS.
The ESRF automated sample changer accepts the Hampton Research CrystalCap Magnetic with an 18
mm mounted cryoloop.
C R Y S T A L C A P

M A G N E T I C
d e s c r i p t i o n
Order Information
Cat. No. Name Description Price
HR4-731 CrystalCap Magnetic with Vial - 10 pack $60.00
HR4-733 CrystalCap Magnetic with Vial - 60 pack $300.00
HR4-902 CrystalCap Magnetic without Vial - 60 pack $145.00
HR4-779 CrystalCap Magnetic ALS without Vial - 60 pack $150.00
HR4-904 CrystalCap Magnetic Vial Vial ONLY - 30 pack $84.00
Fab crystals growing on a thread.
Allen B. Edmundson,
Oklahoma Medical Research Foundation, USA.
9.7 mm
2
1
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5

m
m
12 mm
0.65 mm
9.8 mm
2
1
.
5

m
m
0.65 mm
9.7 mm
12 mm
9.65 mm
CrystalCap Magnetic
CrystalCap Magnetic ALS
C R Y S T A L C A P

M A G N E T I C S Y S T E M S
C R Y S T A L C A P

S Y S T E M S
Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com 122
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Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com
n
Cryocrystallography
n
Magnetic cap

n
Specially designed vial with magnetic ring
n
Cap magnetically attaches to vial
n
Copper jacket to reduce icing on MicroTube
n
Vented vial
n
No threads
n
Vial without magnetic base
a p p l i c a t i o n
f e a t u r e s
n
Cryocrystallography
n
Cap threads to vial

n
Vented cap
n
Magnetic base plate in cap for attachment
to a magnetic base
n
Cap accepts 0.65 mm diameter MicroTube
or Mounted CryoLoop
a p p l i c a t i o n
f e a t u r e s
CrystalCap Copper Magnetic is designed to prevent ice formation
along the copper pin/MicroTube in a cryostream where the flow
is positioned perpendicular, at an angle, or non-collinear to the
MicroTube. The CrystalCap Copper Magnetic is a non-threaded,
1.8 ml (approximate) cryo vial storage container with a specially
engineered cap/pin for cryocrystallography. Each cap is manufac-
tured from an alloy base which allows the cap to be magnetically
secured to a magnet base in the goniometer head. Into the alloy
cap is threaded and bonded a solid, 3 mm diameter copper pin.
The end of the copper pin is machined to an aerodynamic 60 and has a 0.65 mm opening designed
to accept a 10 mm MicroTube and CryoLoop or Mounted CryoLoop snapped at the 10 mm mark
to properly position the CryoLoop in the beam. A ring magnet is molded into the top end of the vial so
that when the cap is positioned in the vial, the ring magnet holds the cap on the vial during cryogenic
storage. The tip of the metal cap is ready to accept a MicroTube and CryoLoop or Mounted CryoLoop.
The vial has two holes in the vial for venting. The sampler pack contains two of each copper pin length
(10, 12, 14, 16, 18, 21, 24 mm). The length is the distance between the top of the copper pin and the
bottom of the CrystalCap Copper Magnetic where contact is made with the magnetic base.
The ESRF automated sample changer accepts the Hampton Research CrystalCap Copper Magnetic
18 mm.
The CrystalCap is a threaded, 1.8 ml cryo vial storage container with
a specially engineered cap for cryocrystallography. Each cap contains
a magnetic base plate which allows the CrystalCap to be magnetically
secured to a magnetic base in the goniometer head. The opposite
end of the CrystalCap is ready to accept a MicroTube and CryoLoop
or Mounted CryoLoop. The CrystalCap lid is vented to allow cryogen
to pass in and out of the container. The CrystalCap is aerodynamically
efficient to minimize ice buildup.
C R Y S T A L C A P

C O P P E R M A G N E T I C
C R Y S T A L C A P

d e s c r i p t i o n
d e s c r i p t i o n
Order Information
Cat. No. Name Description Price
HR4-737 CrystalCap Copper Magnetic - 10 mm with Vial - 30 pack $190.00
HR4-739 CrystalCap Copper Magnetic - 12 mm with Vial - 30 pack $190.00
HR4-741 CrystalCap Copper Magnetic - 14 mm with Vial - 30 pack $190.00
HR4-743 CrystalCap Copper Magnetic - 16 mm with Vial - 30 pack $190.00
HR4-745 CrystalCap Copper Magnetic - 18 mm with Vial - 30 pack $190.00
HR4-747 CrystalCap Copper Magnetic - 21 mm with Vial - 30 pack $190.00
HR4-749 CrystalCap Copper Magnetic - 24 mm with Vial - 30 pack $190.00
HR4-900 CrystalCap Copper Magnetic - 18 mm without Vial - 30 pack $117.00
HR4-904 CrystalCap Magnetic Vial Vial ONLY - 30 pack $84.00
Order Information
Cat. No. Name Description Price

HR4-911 CrystalCap 10 pack $55.00
HR4-913 CrystalCap 60 pack $285.00
HR4-914 CrystalCap Ex, 0.5 - 0.7 mm 30 pack $225.00
123
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C R Y S T A L C A P

S Y S T E M S
n
Cryocrystallography
n
Cap threads to vial

n
Caps are colored to make identification &
organization easier
n
Vented cap
n
Magnetic base plate in cap for attachment to
a magnetic base
n
Cap accepts 0.65 mm diameter MicroTube
and Mounted CryoLoop
a p p l i c a t i o n
f e a t u r e s
The CrystalCap is now available is six different colors: Red,
Orange, Yellow, Green, Blue, and Pink. The vial container
remains clear. Only the cap lid is colored. Same CrystalCap
features are color coded to make identification, storage, and
recovery faster, easier, and less confusing.
The CrystalCap is a threaded, 1.8 ml cryo vial storage container
with a specially engineered cap for cryocrystallography. Each cap
contains a magnetic base plate which allows the CrystalCap to be
magnetically secured to a magnetic base in the goniometer head. The CrystalCap lid is vented to allow
cryogen to pass in and out of the vial container.
C R Y S T A L C A P

- C O L O R E D
d e s c r i p t i o n
Order Information
Cat. No. Name Description Price
HR8-014 CrystalCap Colored - Sampler 60 pack (10 of each color) $300.00
HR8-002 CrystalCap Colored - Red 60 pack $300.00
HR8-004 CrystalCap Colored - Orange 60 pack $300.00
HR8-006 CrystalCap Colored - Yellow 60 pack $300.00
HR8-008 CrystalCap Colored - Green 60 pack $300.00
HR8-010 CrystalCap Colored - Blue 60 pack $300.00
HR8-012 CrystalCap Colored - Pink 60 pack $300.00
CrystalCap Copper is designed to prevent ice formation when using
the MicroTube in a cryostream where the flow is positioned per-
pendicular, at an angle, or non-collinear to the MicroTube. The
CrystalCap Copper uses a 1.8 ml cryo vial storage container and a
cap which contains a sealed, zinc-plated metal core which allows
the CrystalCap Copper to be secured to a magnetic platform on the
goniometer head. Threaded into the metal base is a solid, 3 mm
diameter copper pin. The end of the copper pin is machined to an
aerodynamic 60 and has a 0.65 mm opening to accept ONLY a 10 mm MicroTube and CryoLoop
or Mounted CryoLoop snapped at the 10 mm mark. The CrystalCap Copper lid is vented to allow
cryogen to pass in and out of the container. The length (10, 12, 14, 16, 18, 21, or 24 mm) is the dis-
tance between the top of the copper pin and the bottom of the CrystalCap Copper where contact is
made with the magnetic base.
Order Information
Cat. No. Name Description Price
HR4-969 CrystalCap Copper - 10 mm 30 pack $180.00
HR4-971 CrystalCap Copper - 12 mm 30 pack $180.00
HR4-973 CrystalCap Copper - 14 mm 30 pack $180.00
HR4-665 CrystalCap Copper - 16 mm 30 pack $180.00
HR4-975 CrystalCap Copper - 18 mm 30 pack $180.00
HR4-977 CrystalCap Copper - 21 mm 30 pack $180.00
HR4-979 CrystalCap Copper - 24 mm 30 pack $180.00
n
Cryocrystallography
n
Cap threads to vial
n
Copper jacket to reduce icing on MicroTube
n
Vented cap
n
Zinc plated steel base plate for attachment
to a magnetic base
a p p l i c a t i o n
f e a t u r e s
d e s c r i p t i o n
C R Y S T A L C A P

C O P P E R
124
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C R Y O L O O P S & M I C R O T U B E S
Mounted CryoLoops with 10 micron diameter nylon. These nylon
loops are pre-staked to hollow, stainless steel MicroTubes that
are used to mount, freeze, and secure the crystal during cryocrys-
tallographic procedures and x-ray data collection. The MicroTube
is 24 mm in length and is specially engineered with EasySnap
notches at the 10, 12, 14, 18, and 21 mm measures. To obtain
the desired length of MicroTube, simply snap the MicroTube at
the desired length and stake it to the CrystalCap Magnetic or
CrystalCap Copper Magnetic. These nylon loops show minimal
diffraction, are thin for fast freezing, strong, and aerodynamic. The 10 micron Mounted CryoLoop
shows less background diffraction than the 20 micron Mounted CryoLoop. Please note that the
10 micron Mounted CryoLoops are very flexible due to their small diameter. Most data collection
geometries prefer or require a 22 mm length between the base of the cap and the beam. For a 22 mm
overall length using the CrystalCap, CrystalCap Magnetic and CrystalCap HT, snap the MicroTube at
the second notch from the bottom. Snap the MicroTube at the fifth notch from the bottom (notch
closest to the CryoLoop) and attach to an 18 mm CrystalCap Copper or CrystalCap Copper Magnetic
to create a 22mm length between the base of the cap and the beam.
M O U N T E D C R Y O L O O P

- 1 0 M I C R O N
d e s c r i p t i o n
Order Information
Cat. No. Name Description Price
HR4-993 Mounted CryoLoop - 10 micron Sampler - 25 pack (5 of each size) $67.00
HR4-995 Mounted CryoLoop - 10 micron 0.05 - 0.1 mm - 25 pack $67.00
HR4-997 Mounted CryoLoop - 10 micron 0.1 - 0.2 mm - 25 pack $67.00
HR4-999 Mounted CryoLoop - 10 micron 0.2 - 0.3 mm - 25 pack $67.00
HR4-615 Mounted CryoLoop - 10 micron 0.3 - 0.4 mm - 25 pack $67.00
HR4-617 Mounted CryoLoop - 10 micron 0.4 - 0.5 mm - 25 pack $67.00
n
Cryocrystallography
n
Synthetic CryoLoop on stainless steel
MicroTube

n
MicroTube specially engineered with
EasySnap notches
n
Complete range of loop diameters from
0.025 - 1.0 mm
a p p l i c a t i o n
f e a t u r e s
Mounted CryoLoops with 20 micron diameter nylon. These nylon
loops are pre-staked to hollow, stainless steel MicroTubes that
are used to mount, freeze, and secure the crystal during cryocrys-
tallographic procedures and x-ray data collection. The MicroTube
is 24 mm in length and is specially engineered with EasySnap
notches at the 10, 12, 14, 18, and 21 mm measures. To obtain
the desired length of MicroTube, simply snap the MicroTube at
the desired length and stake it to the CrystalCap Magnetic or
CrystalCap Copper Magnetic. These nylon loops show minimal
diffraction, are thin for fast freezing, strong, and aerodynamic. Most data collection geometries prefer
or require a 22 mm length between the base of the cap and the beam. For a 22 mm overall length
using the CrystalCap, CrystalCap Magnetic and CrystalCap HT, snap the MicroTube at the second
notch from the bottom. Snap the MicroTube at the fifth notch from the bottom (notch closest to the
CryoLoop) and attach to an 18 mm CrystalCap Copper or CrystalCap Copper Magnetic to create a
22 mm length between the base of the cap and the beam.
M O U N T E D C R Y O L O O P

- 2 0 M I C R O N
d e s c r i p t i o n
Order Information
Cat. No. Name Description Price
HR4-953 Mounted CryoLoop - 20 micron Sampler - 30 pack $78.00
(5 of each diameter size 0.1 - 1.0 mm)
HR4-313 Mounted CryoLoop - 20 micron 0.025 - 0.05 mm - 25 pack $62.00
HR4-625 Mounted CryoLoop - 20 micron 0.05 - 0.1 mm - 25 pack $62.00
HR4-955 Mounted CryoLoop - 20 micron 0.1 - 0.2 mm - 25 pack $62.00
HR4-957 Mounted CryoLoop - 20 micron 0.2 - 0.3 mm - 25 pack $62.00
HR4-959 Mounted CryoLoop - 20 micron 0.3 - 0.4 mm - 25 pack $62.00
HR4-961 Mounted CryoLoop - 20 micron 0.4 - 0.5 mm - 25 pack $62.00
HR4-963 Mounted CryoLoop - 20 micron 0.5 - 0.7 mm - 25 pack $62.00
HR4-965 Mounted CryoLoop - 20 micron 0.7 - 1.0 mm - 25 pack $62.00
n
Cryocrystallography
n
Synthetic CryoLoop on stainless steel
MicroTube

n
MicroTube specially engineered with
EasySnap notches
n
Complete range of loop diameters from
0.05 - 0.5 mm
a p p l i c a t i o n
f e a t u r e s
125
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Similar to the Mounted CryoLoop in design and func-
tion, the Adjustable Mounted CryoLoop allows one to
bend and orient the CryoLoop to multiple, unique posi-
tions previously unattainable with typical cryo setups. The
Adjustable Mounted CryoLoop is a 20 micron diameter
synthetic loop mounted inside a malleable stainless steel
sleeve, which in turn, is mounted inside the EasySnap
MicroTube.
Using a specially designed L-shaped tool with a positioning notch, one can manipulate the angle and
orientation of the CryoLoop by bending and adjusting the angle of the malleable stainless steel insert
which holds the CryoLoop. The Adjustable Mounted CryoLoop may be positioned several times. Note:
Repeated manipulation will fatigue the sleeve which can lead to failure of the sleeve. The Adjustable
Mounted CryoLoop is designed to work with the CrystalCap System components.
The MicroTube is 24 mm in length and is specially engineered with EasySnap notches at the 10, 12,
14, 18, and 21 mm measures. To obtain the desired length of MicroTube, simply snap the MicroTube
at the desired length and stake it to the CrystalCap Magnetic or CrystalCap Copper Magnetic.
These nylon loops show minimal diffraction, are thin for fast freezing, strong, and aerodynamic. Most
data collection geometries prefer or require a 22 mm length between the base of the cap and the
beam. For a 22 mm overall length using the CrystalCap, CrystalCap Magnetic and CrystalCap HT, snap
the MicroTube at the second notch from the bottom. Snap the MicroTube at the fifth notch from the
bottom (notch closest to the CryoLoop) and attach to an 18 mm CrystalCap Copper or CrystalCap
Copper Magnetic to create a 22 mm length between the base of the cap and the beam.
A D J U S T A B L E M O U N T E D C R Y O L O O P

d e s c r i p t i o n
Order Information
Cat. No. Name Description Price
HR5-900 Adjustable Mounted CryoLoop Sampler - 30 pack $98.00
(5 of each diameter size 0.1 - 1.0 mm)
HR4-336 Adjustable Mounted CryoLoop 0.025 - 0.05 mm - 25 pack $82.00
HR4-338 Adjustable Mounted CryoLoop 0.05 - 0.1 mm - 25 pack $82.00
HR8-102 Adjustable Mounted CryoLoop 0.1 - 0.2 mm - 25 pack $82.00
HR8-072 Adjustable Mounted CryoLoop 0.2 - 0.3 mm - 25 pack $82.00
HR8-104 Adjustable Mounted CryoLoop 0.3 - 0.4 mm - 25 pack $82.00
HR8-106 Adjustable Mounted CryoLoop 0.4 - 0.5 mm - 25 pack $82.00
HR8-108 Adjustable Mounted CryoLoop 0.5 - 0.7 mm - 25 pack $82.00
HR8-110 Adjustable Mounted CryoLoop 0.7 - 1.0 mm - 25 pack $82.00
HR5-902 Tool for Adjustable each $35.00
Mounted CryoLoop
n
Cryocrystallography
n
Adjustable loop orientation

n
CryoLoops are pre-mounted
n
Synthetic CryoLoop on adjustable stainless
steel MicroTube
n
MicroTube specially engineered with
EasySnap notches
n
Available in 20 micron diameter nylon
n
Customize your CryoLoop orientation
a p p l i c a t i o n
f e a t u r e s
Flower shaped crystal of a hypothetical protein from S.Aureas.
Diana Benetteraj, Nickolay Chirgadze Lab, Clinical Genomics Centre,
University Health Network Max Bell Research Centre, Toronto, Ontario, Canada.
C R Y O L O O P S & M I C R O T U B E S
Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com 126
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The MicroTube is the mount and support for the CryoLoop for crys-
tallographers who prefer to mount their own CryoLoop. It fits into the
pinhole opening on the CrystalCap or CrystalCap Copper (use 10 mm
only) and is secured with epoxy or Super Glue. A CryoLoop is then secured
into the opposite end of the MicroTube. The MicroTube is a laser-cut, 0.65
mm diameter stainless steel hollow tube. They are available in a variety
of lengths to accommodate the wide range of geometries presented by
various x-ray data collection systems. The 18 mm length MicroTube is the
standard length used at most data collection facilities. Try a MicroTube
FitKit pack to determine the best length for your application.
Determining MicroTube Length
Follow the instructions to determine which size works best for you. The following considerations have
been taken in determining the length which is best suited for you:
Most XYZ goniometer heads have 5 mm of Z adjustment. The length of exposed loop stem on a
Mounted CryoLoop is approximately 0.5 - 0.7 mm. When mounted properly, the length of exposed
MicroTube set in a CrystalCap Copper is approximately 1.0 mm.
Recommendations:
Use 18 mm MicroTube with CrystalCap
Use 10 mm MicroTube with CrystalCap Copper 18 mm
CryoLoops in 20 micron diameter nylon. They are used to mount,
freeze, and secure the crystal during cryocrystallographic pro-
cedures and x-ray data collection. These nylon CryoLoops show
minimal diffraction, are thin for fast freezing, strong, and aerody-
namic. Supplied as 10 loops per strip, 6 strips per package. Stake
CryoLoops into MicroTubes using epoxy or Super Glue.
M I C R O T U B E

d e s c r i p t i o n
d e s c r i p t i o n
Order Information
Cat. No. Name Description Price
HR4-928 MicroTube FitKit 6 pack (1 of each size) $25.00
HR4-915 MicroTube - 10 mm 60 pack $59.00
HR4-917 MicroTube - 12 mm 60 pack $59.00
HR4-919 MicroTube - 14 mm 60 pack $59.00
HR4-921 MicroTube - 18 mm 60 pack $59.00
HR4-923 MicroTube - 21 mm 60 pack $59.00
HR4-925 MicroTube - 24 mm 60 pack $59.00
HR4-318 Epoxy Dual Syringe 25 ml tube $10.00
HR4-346 Epoxy & Hardener 35 g tube (2 pack) $9.00
HR4-316 Super Glue 2 g tube $5.00
n
CryoLoop support for CrystalCap
n
Laser-cut steel pin

n
Use to mount CryoLoop to CrystalCap
n
Cryocrystallography
n
Synthetic CryoLoop

n
Complete range of loop diameters
from 0.05 - 1.0 mm
n
20 micron diameter nylon
a p p l i c a t i o n
a p p l i c a t i o n
f e a t u r e s
f e a t u r e s
Order Information
Cat. No. Name Description Price
HR4-941 CryoLoop - 20 micron Sampler - 70 pack (10 of each size) $79.00
HR4-623 CryoLoop - 20 micron 0.05 - 0.1 mm - 60 pack $69.00
HR4-929 CryoLoop - 20 micron 0.1 - 0.2 mm - 60 pack $69.00
HR4-931 CryoLoop - 20 micron 0.2 - 0.3 mm - 60 pack $69.00
HR4-933 CryoLoop - 20 micron 0.3 - 0.4 mm - 60 pack $69.00
HR4-935 CryoLoop - 20 micron 0.4 - 0.5 mm - 60 pack $69.00
HR4-937 CryoLoop - 20 micron 0.5 - 0.7 mm - 60 pack $69.00
HR4-939 CryoLoop - 20 micron 0.7 - 1.0 mm - 60 pack $69.00
Crystal shown being collected using a Hampton Research
CryoLoop.
Picture courtesy of Lisa Edberg. Center for Macromolecular
Crystallography University of Alabama, Birmingham.
C R Y O L O O P S

- 2 0 M I C R O N
127
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CryoLoops are available in 10 and 20 micron diameters. They are
used to mount, freeze, and secure the crystal during cryocrystal-
lographic procedures and x-ray data collection. These synthetic
CryoLoops show minimal diffraction, are thin for fast freezing,
strong, and aerodynamic. Supplied as 10 loops per strip, 6 strips
per package. Stake CryoLoops into MicroTubes using Super Glue
(HR4-316). Please note that the 10 micron CryoLoops are very flex-
ible due to their small diameter.
5 minute, fast drying epoxy for sealing capillary tubes or secur-
ing MicroTubes into CrystalCaps or securing CryoLoops into
MicroTubes. Sets in 5 minutes, can be handled in 15 minutes. Full
bond strength in 1 hour. Requires no heat.
Super Glue for securing CryoLoops into MicroTubes.
C R Y O L O O P S

- 1 0 M I C R O N
E P O X Y
S U P E R G L U E
d e s c r i p t i o n
d e s c r i p t i o n
d e s c r i p t i o n
n
Cryocrystallography
n
Synthetic CryoLoop

n
Complete range of loop diameters
from 0.05 - 0.5 mm
n
10 micron diameter nylon
n
Sealing capillary tubes
n
Securing MicroTubes into CrystalCaps

n
Securing CryoLoops into MicroTubes
n
Staking CryoLoops to MicroTubes
a p p l i c a t i o n
a p p l i c a t i o n s
a p p l i c a t i o n
f e a t u r e s
Order Information
Cat. No. Name Description Price
HR4-981 CryoLoop - 10 micron Sampler - 50 pack (10 of each size) $69.00
HR4-983 CryoLoop - 10 micron 0.05 - 0.1 mm - 60 pack $79.00
HR4-985 CryoLoop - 10 micron 0.1 - 0.2 mm - 60 pack $79.00
HR4-987 CryoLoop - 10 micron 0.2 - 0.3 mm - 60 pack $79.00
HR4-989 CryoLoop - 10 micron 0.3 - 0.4 mm - 60 pack $79.00
HR4-991 CryoLoop - 10 micron 0.4 - 0.5 mm - 60 pack $79.00
Crystal shown being collected using a Hampton Research CryoLoop.
Picture courtesy of Lisa Edberg. Center for Macromolecular Crystallography University of Alabama, Birmingham.
Order Information
Cat. No. Name Description Price
HR4-318 Epoxy Dual Syringe 25 ml tube $10.00
HR4-346 Epoxy & Hardener 35 g tube (2 pack) $9.00
Order Information
Cat. No. Name Description Price
HR4-316 Super Glue 2 g tube $5.00
C R Y O T O O L S
128
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The CryoTong is a tool used to manually transfer a crystal mounted
on a CrystalCap from liquid nitrogen to a magnetic base in a
goniometer head positioned in a cryogenic stream, and then back
to liquid nitrogen. The one-piece, compact CryoTong is available
in two lengths. The standard CryoTong is approximately 110 mm
in length from the edge of the tool head to the end of the handle.
The artery clamp style maintains the CryoTong in the closed posi-
tion until the clamp is squeezed, which opens the opposing heads.
The heads are non-magnetic stainless steel. The handle is magnetic stainless steel. The inside of the
head is machined to closely surround the CrystalCap with loop and crystal in place. A small retaining lip
is machined into the lower portion of the head to prevent the CrystalCap from slipping out when the
tool is in the closed position. The CryoTong can maintain the temperature of the crystal at -160C for
up to 30 seconds during room temperature crystal transfers. The CryoTong is available in seven differ-
ent sizes to fit 10 to 24 mm pin heights. Each size is designed to fit any of the the CrystalCap systems.
Choose the proper CryoTong size based upon the pin length required by the configuration of the x-ray
data collection hardware used. The 18 mm CryoTong (HR4-637) is to be used with the CrystalCap HT
systems and any CrystalCap configured to 18 mm.
C R Y O T O N G

- S T A N D A R D
d e s c r i p t i o n
n
Crystal transfer under cryo temperature
n
Artery clamp style closure

n
30 seconds of cryo temperature
n
7 different sizes
n
110 mm length
a p p l i c a t i o n
f e a t u r e s
Order Information
Cat. No. Name Description Price
HR4-631 CryoTong - 10 mm each $55.00
HR4-633 CryoTong - 12 mm each $55.00
HR4-635 CryoTong - 14 mm each $55.00
HR4-667 CryoTong - 16 mm* each $55.00
HR4-637 CryoTong - 18 mm each $55.00
HR4-639 CryoTong - 21 mm each $55.00
HR4-641 CryoTong - 24 mm each $55.00
*The 16 mm CryoTong only works with the CrystalCap Copper 16 mm and the CrystalCap Copper
Magnetic 16 mm.
Valentines Day crystal.
Igor Nederlof, Key Drug Prototyping BV, The Netherlands.
129
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The CryoTong is a tool used to manually transfer a crystal mounted
on a CrystalCap from liquid nitrogen to a magnetic base in a
goniometer head positioned in a cryogenic stream, and then back
to liquid nitrogen. The one-piece, compact CryoTong is available
in two lengths. The long CryoTong is approximately 180 mm in
overall length. The artery clamp style maintains the CryoTong
in the closed position until the clamp is squeezed, which opens
the opposing heads. The heads are non-magnetic stainless steel.
The handle is magnetic stainless steel. The inside of the head is
machined to closely surround the CrystalCap with loop and crystal in place. A small retaining lip is
machined into the lower portion of the head to prevent the CrystalCap from slipping out when the
tool is in the closed position. The CryoTong can maintain the temperature of the crystal at -160C for
up to 30 seconds during room temperature crystal transfers. The CryoTong is available in seven dif-
ferent sizes to fit 10 to 24 mm pin heights. Each size is designed to fit all CrystalCap systems. Choose
the proper CryoTong size based upon the pin length required by the configuration of the x-ray data
collection hardware used. The 18 mm CryoTong (HR5-112) is to be used with the CrystalCap HT and
any other CrystalCap configured for 18 mm.
HR5-114 Long CryoTong 18 mm, 180 features the head in alignment with the handle of the
CryoTong.
HR5-113 Long CryoTong 18 mm, 45 features the head at a 45 angle to the handle of the CryoTong.
C R Y O T O N G

- L O N G
d e s c r i p t i o n
n
Crystal transfer under cryo temperature
n
Artery clamp style closure

n
30 seconds of cryo temperature
n
7 different sizes
n
180 mm length
a p p l i c a t i o n
f e a t u r e s
Order Information
Cat. No. Name Description Price
HR5-104 Long CryoTong - 10 mm each $61.00
HR5-106 Long CryoTong - 12 mm each $61.00
HR5-108 Long CryoTong - 14 mm each $61.00
HR5-110 Long CryoTong - 16 mm* each $61.00
HR5-112 Long CryoTong - 18 mm each $61.00
HR5-113 Long CryoTong - 18 mm, 45 each $61.00
HR5-114 Long CryoTong - 18 mm, 180 each $61.00
HR5-118 Long CryoTong - 21 mm each $61.00
HR5-120 Long CryoTong - 24 mm each $61.00
*The 16 mm Long CryoTong only works with the CrystalCap Copper 16 mm and the CrystalCap Copper
Magnetic 16 mm.
Christmas tree crystals.
Heidi Roth, Department of Structural Biology,
University of Wuerzburg, Germany.
Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com 130
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C R Y O T O O L S
The Long CrystalWand is a chrome-plated steel wand, 9.6 mm in
diameter with a magnet molded inside one end. The magnet is
designed to fit the CrystalCap and CrystalCap Copper. The Long
Crystal Wand makes it easy to manipulate, seed, mount, store, and
handle crystals with the CrystalCap System. It has a plastic handle
and is available in two styles (no tab and with tab) and two lengths
(standard 130 mm and long 205 mm overall length).
Note: Long CrystalWand is not to be used with the CrystalCap
Magnetic or CrystalCap Copper Magnetic or CrystalCap HT. Use the CrystalWand Magnetic for these
products.
The CrystalWand Magnetic is designed to be used exclusively
with the CrystalCap Magnetic, CrystalCap Copper Magnetic, and
CrystalCap HT systems during transfer of the caps from the vial
to the goniometer head and from the goniometer to the vial.
The 6 1/2 (165 mm) long chrome plated steel wand features a
plastic housing enclosing a spring tensioned plunger that when
depressed, moves a non-magnetic steel platform away from the
magnet housed in the end of the wand. This causes the steel
CrystalCap to detach from the wand and be placed readily into the
vial or the CryoTong. When the platform is retracted, the wand securely holds the steel CrystalCap
Magnetic. The CrystalWand Magnetic 45 (McMiken Tool) offers the same functionality but is bent at
a 45 angle at approximately 2.5 cm from the magnet end.
C R Y S T A L WA N D

- L O N G
C R Y S T A L WA N D

M A G N E T I C
d e s c r i p t i o n
d e s c r i p t i o n
n
CrystalCap and CrystalCap Copper handling
tool
n
CrystalCap Magnetic & CrystalCap Copper
Magnetic handling tool
a p p l i c a t i o n
a p p l i c a t i o n
Order Information
Cat. No. Name Description Price
HR4-600 Long CrystalWand no Tab, 205 mm each $46.00
HR4-602 Long CrystalWand with Tab, 205 mm each $46.00
Order Information
Cat. No. Name Description Price
HR4-729 CrystalWand Magnetic, Straight each $61.00
HR4-315 CrystalWand Magnetic, 45 (McMiken Tool) each $56.00
The CrystalWand is a chrome-plated steel wand, 9.6 mm in
diameter with a magnet molded inside one end. The magnet is
designed to fit the CrystalCap and CrystalCap Copper. The Crystal
Wand makes it easy to manipulate, seed, mount, store, and handle
crystals with the CrystalCap system. The CrystalWand is available
in two styles (no tab and with tab) and two lengths (standard 130
mm and long 205 mm overall length). The long CrystalWand has
a plastic handle.
Note: CrystalWand is not to be used with the CrystalCap Magnetic
or CrystalCap Copper Magnetic or CrystalCap HT. Use the
CrystalWand Magnetic for these products.
C R Y S T A L WA N D

- S T A N D A R D
d e s c r i p t i o n
n
CrystalCap & CrystalCap Copper
handling tool
a p p l i c a t i o n
Order Information
Cat. No. Name Description Price
HR4-951 CrystalWand no Tab, 130 mm each $36.00
HR4-619 CrystalWand with Tab, 130 mm each $36.00
CrystalWand With Tab
CrystalWand No Tab
131
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The Vial Clamp - Straight is a chrome plated, hemostat style tool.
It has a tip shaped to hold the storage vial of all the CrystalCap
systems straight when the clamp is closed. The clamp can be
locked using the hemostat style closure. This clamp makes it easy
to dip the storage vial into liquid nitrogen for crystal storage. The
complete length of the clamp is straight, with an overall length of
195 mm. When the vial is placed in the clamp, the length of it is
positioned perpendicular, 90 to the length of the clamp.
d e s c r i p t i o n
n
Vial support and manipulation
n
Straight end tip
n
Hemostat style closure
a p p l i c a t i o n
Order Information
Cat. No. Name Description Price
HR4-670 Vial Clamp - Straight each $49.00
f e a t u r e s
V I A L C L A M P

- S T R A I G H T
The Vial Clamp - Curved is a chrome plated, hemostat style tool.
It has a tip shaped to hold the storage vial of all the CrystalCap
systems at an angle when the clamp is closed. The clamp can be
locked using the hemostat style closure. The end of the clamp,
where the vial is held, is curved at either a 45/135 or 110/ 70
angle. When the vial is placed in the clamp, the length of it is
positioned to the clamp at either of those same angles. The overall
length of the clamp is 195 mm.
The 7 1/2 (190 mm) long, 15 ml Tube Clamp is useful for hold-
ing and manipulating 15 ml screw top centrifuge tubes during
cryocrystallographic procedures involving liquid propane and
liquid nitrogen. The jaws of the clamp are designed to fit the out-
side diameter (16 mm) of most 15 ml screw top centrifuge tubes.
When the vial is placed in the clamp, the length of it is positioned
to the clamp at either a 45 or 135 angle, depending upon how
one holds the clamp.
d e s c r i p t i o n
d e s c r i p t i o n
n
Vial support and manipulation
n
Curved end tip
n
Hemostat style closure
n
15 ml vial support & manipulation
n
Curved end tip
n
Hemostat style closure
a p p l i c a t i o n
a p p l i c a t i o n
Order Information
Cat. No. Name Description Price
HR4-671 Vial Clamp - Curved 45/135 each $49.00
HR4-672 Vial Clamp - Curved 110/70 each $75.00
Order Information
Cat. No. Name Description Price
HR4-727 Tube Clamp each $61.00
f e a t u r e s
f e a t u r e s
V I A L C L A M P

- C U R V E D
T U B E C L A M P

C R Y O T O O L S
Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com 132 Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com
CrystalCap Holders are convenient stands for holding all CrystalCap
systems in liquid nitrogen dewars during crystal cryogenic proce-
dures. Available in two sizes. Both holders feature spring-clips to
secure the CrystalCap Vials in the holder. Both stands have 3"
of adjustable Z (height) which allows one to raise or lower the
CrystalCaps to the appropriate height, depending upon the liquid
nitrogen level in the dewar. Each holder is supplied with a socket
driver for height adjustment.
500 ml version holds five CrystalCaps, two 15 ml vials (for liquid
propane procedures) and fits into the 500 ml dewar. 1,000 ml ver-
sion holds eight CrystalCaps, two 15 ml vials (for liquid propane
procedures) and fits into the 1,000 ml dewar.
The HR4-705 is approximately 98 mm wide (measured at corners of
hexagon)/92 mm wide (measured at flats of hexagon) by approxi-
mately 120 mm tall.
d e s c r i p t i o n
n
CrystalCap support inside stainless steel
dewars
n
Adjustable height
n
Supports up to 8 CrystalCaps
n
15 ml vial position for liquid propane
n
500 and 1,000 ml versions
a p p l i c a t i o n
f e a t u r e s
C R Y S T A L C A P

H O L D E R
Order Information
Cat. No. Name Description Price
HR4-707 CrystalCap Holder - 500 ml each $109.00
HR4-705 CrystalCap Holder - 1,000 ml each $109.00
HR4-706 Replacement Height Adjustment Tool each $15.00
Protein crystals.
Ivana Tomcova, University of South Bohemia & Academy of Science
of the Czech Republic, Nove Hrady, Czech Republic.
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C A N E S , S L E E V E S & C O D E R S
133
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Aluminum CryoCanes firmly hold CrystalCap, CrystalCap Copper,
CrystalCap Magnetic, and CrystalCap Copper Magnetic vials for
storage in dewar-type, liquid nitrogen freezers. The 5 vial CryoCane
offers base tabs for proper CrystalCap seating whereas the 6 vial
CryoCane offers free alignment. Length of 5 vial version: 11 5/16
(28.7 cm). Length of 6 vial version: 11 13/16 (30 cm).
Colored aluminum tabs slip into the ends of CryoCane holders to
identify a set of vials, a researchers work, or sample. Flat writing
surface allows for additional identification. Available in five differ-
ent colors.
d e s c r i p t i o n
d e s c r i p t i o n
n
Organized storage of CrystalCaps
n
2 styles: 5 or 6 vial holding canes
n
Made of aluminum
n
Compatible with the CrystalCap systems
n
Identify & organize CryoCanes
n
Compatible with the CrystalCap systems
n
Made of aluminum
n
5 colors to choose from
a p p l i c a t i o n
a p p l i c a t i o n
Order Information
Cat. No. Name Description Price
HR4-709 CryoCane 5 Vial Holder 12 pack $24.00
HR4-711 CryoCane 6 Vial Holder 12 pack $26.00
f e a t u r e s
f e a t u r e s
C R Y O C A N E
C R Y O C A N E C O L O R C O D E R S
Order Information
Cat. No. Name Description Price
HR4-722 CryoCane Color Coder - Sampler 60 pack (12 of each color) $50.00
HR4-713 CryoCane Color Coder - White 12 pack $10.00
HR4-715 CryoCane Color Coder - Yellow 12 pack $10.00
HR4-717 CryoCane Color Coder - Blue 12 pack $10.00
HR4-719 CryoCane Color Coder - Green 12 pack $10.00
HR4-721 CryoCane Color Coder - Red 12 pack $10.00
Clear polyvinyl sleeve encloses CrystalCap on a CryoCane Holder
for extra security during handling, transport, and storage. Will not
become brittle during freezing or thawing. Unlike cardboard, will
not shred, tear, or fall to pieces after repeated use. 10 13/16 length
(27.5 cm).
d e s c r i p t i o n
n
Use with CryoCanes
n
Will not become brittle
n
Made of polyvinyl
n
Compatible with the CrystalCap systems
a p p l i c a t i o n
f e a t u r e s
C R Y O S L E E V E
Order Information
Cat. No. Name Description Price
HR4-708 CryoSleeve 12 pack $18.00
D E WA R S
Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com 134 Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com
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Low form, glass, shallow dewar for use with materials ranging from
dry ice/acetone to liquid nitrogen. This 600 ml full base hemi-
spherical bottom dewar is a low form with full aluminum protective
coating. Low sidewall allows easier access to cryogen.
Dimensions: ID 140 mm, inside depth 65 mm, OD 165 mm,
total height 95 mm. Accepts 500 ml flask. Manufactured by Pope
Scientific.
For optimum results, temper the Dewar per instructions included with each dewar. Refer to The Care
and Use of Pope Dewar Flasks.
d e s c r i p t i o n
n
Cryogenic procedures
n
Low form, shallow 600 ml base
n
Low sidewall for easier access
n
Manufactured for service of liquid nitrogen
a p p l i c a t i o n
f e a t u r e s
Order Information
Cat. No. Name Description Price
HR5-102 Low Form Dewar each $268.00
G L A S S D E WA R
Unbreakable, 304 stainless steel construction dewars for cryocrys-
tallography. Convenient for the flash freezing of crystals, these
dewars store and transport low temperature liquid gases such as
liquid nitrogen and can withstand temperatures from -196C to
204C. The dewars are impervious to vibrations. In comparison
tests, these stainless steel dewars outperform common metal
dewars by 20% and perform comparable to glass modules. Only
the 1,000 ml dewar comes with a stainless steel carrying handle.
Each dewar is supplied with a stainless steel cover. Covers are also
available separately.
500 ml stainless steel dewar flask (2.5ID, 3.4OD, 7.0 depth, 8 height) - right in the picture. 1,000 ml
stainless steel dewar flask (3.9ID, 4.8 OD, 6.2 depth, 6.9 height) - left in the picture.
d e s c r i p t i o n
n
Cryogenic procedures
n
Made of 304 stainless steel
n
Available in 500 ml and 1,000 ml sizes
n
Compatible with CrystalCap Holders
a p p l i c a t i o n
f e a t u r e s
Order Information
Cat. No. Name Description Price
HR4-695 Dewar Flask - 500 ml with cover each $90.00
HR4-697 Dewar Cover - 500 ml each $20.00
HR4-699 Dewar Flask - 1,000 ml with cover each $120.00
HR4-701 Dewar Cover -1,000 ml each $20.00
S T A I N L E S S S T E E L D E WA R
135
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Spearlab Cryogenic Foam Dewar (800 ml) - Standard: The standard
foam dewar shape is circular, with a protruding handle, so that it
resembles a large teardrop. Each dewar is supplied with a matching
foam lid to insulate the contents from ambient air. Dimensions of
the cylindrical cavity in this vessel are 5.8" in diameter by 2.8" deep,
so that it easily holds 800 ml of liquid nitrogen.
Spearlab Cryogenic Foam Dewar (2 liter) - Tall: The tall foam dewar
shape is a tapered octagon on the outside with a cylindrical inte-
rior. The tapered octagon features a wide, stable base. Each dewar
is supplied with a matching foam lid to insulate the contents from
ambient air. Dimensions of the cylindrical cavity in this vessel are
3.5" in diameter by 12.5" deep, so that it holds approximately 1,970
ml of liquid nitrogen. The outside dimensions of the tall dewar are
15.25" tall with a 6.25" wide top and a 7.75" wide base (measured
on the flats of the octagon). The cover is 6.5" in diameter and 0.5"
thick.
The patent pending design of the Spearlab Cryogenic Foam Dewar
makes it easy to handle and safer to use than a traditional low
profile glass dewar. Also, because of its lower thermal mass, a foam
vessel will cause less boil off when it is filled with liquid nitrogen.
Additionally, the dewar will accumulate less frost during regular
use. The end result is that less liquid nitrogen is consumed.
d e s c r i p t i o n
n
Cryocrystallography
n
Each dewar is supplied with a lid
n
Proprietary foam construction
n
Standard (800 ml) and Tall (2 liter)
a p p l i c a t i o n
f e a t u r e s
Order Information
Cat. No. Name Description Price
HR4-673 Cryogenic Foam Dewar (800 ml) - Standard each $100.00
HR4-675 Cryogenic Foam Dewar (2 liter) - Tall each $160.00
F O A M D E WA R
Crystal of a protein-DNA complex that grew over
wells containing 5% PEG 8000.
Dr. Christopher Davies, Medical University of South Carolina, USA.
HR4-673
HR4-675
c a p i l l a r y m o u n t s & s u p p l i e s
Crystal flakes - Crystals from a RNA duplex.
Vanessa Delfosse and Dr Claudine Mayer,
Laboratoire de Recherche Molculaire sur les Antibiotiques, Paris, France.
T A B L E O F C O N T E N T S
138 - 140 c a p i l l a r i e s
140 g l a s s f i b e r s
141 w i c k s
142 c a p i l l a r y s e a l a n t s
143 - 144 w a x e s & c l a y
145 m a g n e t i c m o u n t s & s u p p o r t s
P A G E S
138
n
X-ray data collection
n
Liquid-liquid diffusion crystallization
n
Gel acupuncture crystallization
n
3 glass types
n
Thin walled
C A P I L L A R I E S
Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com
a p p l i c a t i o n s
f e a t u r e s
Crystal Clear Glass and Quartz capillaries that
are extremely thin walled (approximately 10
microns wall thickness). The length of the capil-
lary has a well defined diameter, with one end
having a funnel shape and the other end closed.
Capillaries have a wall thickness of 0.01 mm and
an overall length of 80 mm. Capillaries are avail-
able in a wide range of outside diameters from
0.1 to 2.0 mm (5.0 mm for quartz only). The cap-
illaries are designed to mount, hold, and store
small molecule and biological macromolecular
crystals for x-ray data collection. Capillaries can
also be used for crystal density measurements
and crystal growth experiments. The capillaries
can be sealed tightly against moisture and gases
using wax, epoxy, or other sealing materials.
In determining what glass or quartz capillary
is right for you, please refer to the Linear
Absorption Coefficient cm
-1
table. This table
indicates the amount of radiation that is absorbed
by the capillary during x-ray data collection.
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d e s c r i p t i o n
Glass Type
10
50
Quartz
CuK L Radiation
126.0
71.0
75.8
MoK L Radiation
14.7
7.35
8.2
Softening Temp.
708C
815C
1,730C
Linear Absorption Coefficient cm
-1
how t o: mount i ng cryst als usi ng capi llari es
Step 1 - Choosing a crystal
Locate the crystal of
interest to be mounted
in the capillary. Measure
the size of crystal. This
will determine what
diameter capillary to use.
Step 2 - Cutting the capillary
Select the size of capillary. Score the capillary near
the sealed end using the Capillary Cutting Stone.
Snap the capillary at the score mark. A clean cut
every time!
Step 3 - Make a slug
On the funnel end of the
capillary attach a flexible
adaptor to use a syringe
or pipet. Suck up a small
amount of mother liquor.
This will be your mother
liquor slug to prevent the
crystal from drying out during data collection.
Step 4 - Obtaining the crystal
Remove the capillary
from the drop and
place an air gap into
the capillary. This will
separate the slug from
the crystal.
Place the capillary back into the drop and suck up
the crystal. Try not to obtain too much mother
liquor while picking up the crystal.
Step 5 - Removing excess mother liquor
Using either the Paper Wicks or the MicroWick,
remove any excess mother liquor from around the
crystal. There should only be a minimal amount
of mother liquor surrounding the crystal. This will
help reduce background scattering and reduce
crystal slippage.
Step 6 - Sealing one end
Using either Beeswax or Capillary Wax, seal off the
open end of the capillary. Do not overheat the wax
for this will move or even damage the crystal.
Step 7 - Sealing the other end
Trim off the funnel part of the capillary using the
Capillary Cutting Stone as described in Step 2.
Finally, seal off the open end of the capillary as
described in Step 6.
Using Mounting Clay or an Adjustable Crystal
Mount, and a Brass Pin, place the capillary contain-
ing your crystal onto the goniometer head. You are
now ready to collect data.
139
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Order Information
Cat. No. Name Description Price
HR6-152 Special Glass 10 Capillary 0.1 mm - 25 pack $65.00
HR6-154 Special Glass 10 Capillary 0.2 mm - 25 pack $65.00
HR6-156 Special Glass 10 Capillary 0.3 mm - 25 pack $65.00
HR6-158 Special Glass 10 Capillary 0.4 mm - 25 pack $65.00
HR6-160 Special Glass 10 Capillary 0.5 mm - 25 pack $65.00
HR6-162 Special Glass 10 Capillary 0.6 mm - 25 pack $65.00
HR6-164 Special Glass 10 Capillary 0.7 mm - 25 pack $65.00
HR6-166 Special Glass 10 Capillary 0.8 mm - 25 pack $65.00
HR6-168 Special Glass 10 Capillary 0.9 mm - 25 pack $65.00
HR6-170 Special Glass 10 Capillary 1.0 mm - 25 pack $65.00
HR6-172 Special Glass 10 Capillary 1.5 mm - 25 pack $65.00
HR6-174 Special Glass 10 Capillary 2.0 mm - 25 pack $85.00
Order Information
Cat. No. Name Description Price
HR6-104 Glass Number 50 Capillary 0.1 mm - 25 pack $78.00
HR6-106 Glass Number 50 Capillary 0.2 mm - 25 pack $78.00
HR6-108 Glass Number 50 Capillary 0.3 mm - 25 pack $78.00
HR6-110 Glass Number 50 Capillary 0.4 mm - 25 pack $78.00
HR6-112 Glass Number 50 Capillary 0.5 mm - 25 pack $78.00
HR6-114 Glass Number 50 Capillary 0.6 mm - 25 pack $78.00
HR6-116 Glass Number 50 Capillary 0.7 mm - 25 pack $78.00
HR6-118 Glass Number 50 Capillary 0.8 mm - 25 pack $78.00
HR6-120 Glass Number 50 Capillary 0.9 mm - 25 pack $78.00
HR6-122 Glass Number 50 Capillary 1.0 mm - 25 pack $78.00
HR6-124 Glass Number 50 Capillary 1.5 mm - 25 pack $88.00
HR6-126 Glass Number 50 Capillary 2.0 mm - 25 pack $98.00
Order Information
Cat. No. Name Description Price
HR6-128 Quartz Capillary 0.1 mm - 25 pack $118.00
HR6-130 Quartz Capillary 0.2 mm - 25 pack $118.00
HR6-132 Quartz Capillary 0.3 mm - 25 pack $118.00
HR6-134 Quartz Capillary 0.4 mm - 25 pack $118.00
HR6-136 Quartz Capillary 0.5 mm - 25 pack $118.00
HR6-138 Quartz Capillary 0.6 mm - 25 pack $118.00
HR6-140 Quartz Capillary 0.7 mm - 25 pack $118.00
HR6-142 Quartz Capillary 0.8 mm - 25 pack $118.00
HR6-144 Quartz Capillary 0.9 mm - 25 pack $118.00
HR6-146 Quartz Capillary 1.0 mm - 25 pack $118.00
HR6-148 Quartz Capillary 1.5 mm - 25 pack $118.00
HR6-150 Quartz Capillary 2.0 mm - 25 pack $128.00
HR6-151 Quartz Capillary 2.5 mm - 15 pack $128.00
HR6-175 Quartz Capillary 3.0 mm - 15 pack $240.00
HR6-177 Quartz Capillary 4.0 mm - 5 pack $240.00
HR6-179 Quartz Capillary 5.0 mm - 5 pack $285.00
s p e c i a l g l a s s 1 0 ( S O D A L I ME G L A S S )
g l a s s n u m b e r 5 0 ( B O R O S I L I C AT E )
q u a r t z
C A P I L L A R I E S
G L A S S F I B E R S
Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com
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a.) b.) c.)
40 mm length glass fibers for small molecule crystal mounts.
Available in 0.1 - 0.3 mm and 0.3 - 0.5 mm diameter glass. Simple
to use. Glue the glass fiber to a brass specimen pin. Adhesives for
attaching glass fibers to the brass pins can be found in Capillary
Essentials. Add a very small amount of the adhesive of choice to the
tip of the glass fiber. Carefully bring the fiber into solid contact with
the crystal face and leave in this orientation long enough for the
glue to set firmly. Position onto goniometer head and collect data.
These small (25 mm x 25 mm x 0.65 mm) cutting stones are used
to make clean cuts of glass and quartz capillaries. Use the edge
of the stone to etch the capillary then gently snap the capillary in
two. This is useful for cutting the tips off closed capillaries used for
mounting single crystals for x-ray diffraction analysis or for cutting
capillaries to the desired length.
n
Crystal mounts for small molecules
n
Two sizes: 0.1 - 0.3 mm & 0.3 - 0.5 mm
n
Length: 40 mm
n
The easiest way to cut capillaries
n
Score and snap
a p p l i c a t i o n
a p p l i c a t i o n
f e a t u r e s
f e a t u r e s
d e s c r i p t i o n
d e s c r i p t i o n
C A P I L L A R Y C U T T I N G S T O N E

Order Information
Cat. No. Name Description Price
HR8-030 Glass Fiber 0.1 - 0.3 mm diameter - 25 pack $15.00
HR8-032 Glass Fiber 0.3 - 0.5 mm diameter - 25 pack $15.00
Order Information
Cat. No. Name Description Price
HR4-334 Capillary Cutting Stone each $15.00
140
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W I C K S
141
Sterile Paper Wicks are available in a variety of sizes. Smaller
diameter wicks (in 25 mm & 55 mm lengths) are useful for
wicking away mother liquor from within capillary tubes and
CryoLoops while mounting crystals. Larger diameter wicks (in 25
mm & 55 mm lengths) are useful for removing stray/excess solvent
from crystallization setups.
n
Remove excess mother liquor from inside
capillaries and CryoLoops
n
Remove stray drops
a p p l i c a t i o n s
d e s c r i p t i o n
P A P E R W I C K S

Order Information
Cat. No. Name Description Price
HR4-110 Assorted Paper Wicks - 25 mm XXX Fine - X Fine - 200 wicks $15.00
HR4-122 Paper Wicks - 25 mm XXX Fine - 200 wicks $15.00
HR4-112 Paper Wicks - 25 mm XX Fine - 200 wicks $15.00
HR4-114 Paper Wicks - 25 mm X Fine - 200 wicks $15.00
HR4-116 Paper Wicks - 25 mm Fine - 200 wicks $15.00
HR4-211 Paper Wicks - 55 mm X Fine Long - 100 wicks $15.00
HR4-213 Paper Wicks - 55 mm Medium Long - 100 wicks $15.00
MicroWick is useful for removing mother liquor from inside capil-
laries during crystal mounting. MicroWick is a non-metallic syringe
needle used for filling and removing liquid from capillaries and
micropipettes. MicroWick features a 70 mm tip, 0.164 mm OD,
0.1 mm ID (34 gauge) tip which will fit into most capillaries and
micropipettes with an internal diameter of 0.2 mm or larger. The
MicroWick needle is constructed from a combination of plastic
and fused silica - no metal components are used. The MicroWick
tip elasticity is sturdy and flexible though not unbreakable (much more flexible than glass!). Moderate
bending will not block or damage the needle. The MicroWick luer fitting allows easy coupling to
syringes and syringe filters. Each MicroWick contains one needle and one 1 ml syringe. Crystallization
and capillary accessories sold separately.
n
Remove excess mother liquor from inside
capillaries
a p p l i c a t i o n
d e s c r i p t i o n
M I C R O W I C K

Order Information
Cat. No. Name Description Price
HR4-330 MicroWick each $31.00
C A P I L L A R Y E S S E N T I A L S
Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com
C A P I L L A R Y S E A L A N T S
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142
5 minute, fast drying epoxy for sealing capillary tubes or stak-
ing MicroTubes into CrystalCaps. Sets in 5 minutes, can be
handled in 15 minutes. Full bond strength in 1 hour. Requires no
heat!
Old school cement for sealing glass capillaries when mounting
crystals. Requires no heat!
Super glue for staking glass fibers to brass specimen pins, crystal
mounting, and sticking fingers together.
n
Sealing capillaries
n
Staking MicroTubes into CrystalCaps
n
Mount crystals to glass fibers
n
Sealing capillaries
n
Mount crystals to glass fibers
n
Staking glass fibers to brass specimen pins
n
Staking CryoLoops to MicroTubes
a p p l i c a t i o n s
a p p l i c a t i o n s
a p p l i c a t i o n s
d e s c r i p t i o n
d e s c r i p t i o n
d e s c r i p t i o n
E P O X Y
D U C O

C E M E N T
S U P E R G L U E
Order Information
Cat. No. Name Description Price
HR4-320 Duco Cement 29 ml tube $7.00
Order Information
Cat. No. Name Description Price
HR4-316 Super Glue 2 g tube $5.00
Order Information
Cat. No. Name Description Price
HR4-318 Epoxy Dual Syringe 25 ml tube $10.00
HR4-346 Epoxy & Hardener 35 g tube (2 pack) $9.00
n
Sealing capillaries
n
Keep bees busy
a p p l i c a t i o n s
n
Sealing capillaries
a p p l i c a t i o n
n
Sealing capillaries
a p p l i c a t i o n
Order Information
Cat. No. Name Description Price
HR4-312 Beeswax Stick 7 sticks $30.00
Pure, natural, and soft Beeswax - golden in color. Beeswax has a
melting point of 63C/146F. Beeswax comes in seven easy to peel
wrapped sticks making it very convenient to handle. Very useful
for sealing the ends of capillary tubes when mounting crystals.
B E E S WA X

d e s c r i p t i o n
A hard, brittle wax orange in color useful for sealing crystals in
glass capillaries. Non-malleable. Melt with soldering iron or other
heat device and apply to glass or quartz capillaries. When applied
properly, Capillary Wax will provide an airtight seal. Also referred
to as sticky wax.
C A P I L L A R Y WA X

d e s c r i p t i o n
The Wax Pen is a hot pen, powered by two AA batteries
(supplied). Pressing a switch heats the tips of the pen to a tem-
perature that melts Beeswax and Capillary Wax which are used
for sealing capillaries.
Replacement tips are available in a pack of three different shaped
tips (straight loop, straight flat, and curved loop).
WA X P E N

d e s c r i p t i o n
Order Information
Cat. No. Name Description Price
HR4-328 Capillary Wax 40 g pack (approximate) $25.00
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WA X E S & C L A Y
143
Order Information
Cat. No. Name Description Price
HR4-342 Wax Pen each $50.00
HR4-344 Wax Pen Replacement Tips 3 tip pack $21.00
C A P I L L A R Y E S S E N T I A L S
Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com
C A P I L L A R Y E S S E N T I A L S WA X E S & C L A Y
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Adhesive, malleable, non-hardening clays have many uses in the
crystallization lab. Useful for holding capillaries during crystal
mounting as well as, stabilizing the capillary in the goniometer
head. Useful for propping open the lids of crystallization plates.
Great for color coding experiments, capillaries, and Linbro

crys-
tallization plates. Each 4 oz brick of clay is individually wrapped.
F O U R C O L O R M O U N T I N G C L A Y
n
Stabilizing the capillary on the goniometer
head or brass specimen pin
n
Color code capillary mounts
a p p l i c a t i o n
d e s c r i p t i o n
f e a t u r e s
Order Information
Cat. No. Name Description Price
HR4-326 Four Color Mounting Clay 16 oz pack $12.00
(4 oz each blue/yellow/red/green)
Soft, malleable, cylindrical wax ropes with similar properties as
mounting clay. Uses include propping open the lids of Linbro

crystallization plates, supporting small items, and stabilizing
capillaries.
R E D S T I C K Y WA X
n
Stabilizing the capillary on the goniometer
head or brass specimen pin
n
Prop open lids of Linbro

plates
n
Make bendable buddies
a p p l i c a t i o n s
d e s c r i p t i o n
Order Information
Cat. No. Name Description Price
HR4-310 Red Sticky Wax 44 wax ropes $20.00
C A P I L L A R Y E S S E N T I A L S
Vvn protein with DNA "diamond clock".
Lucy Doudeva, Hanna Yuan's lab, Academia Sinica, Taiwan.
d e s c r i p t i o n
n
Adjustable, magnetic crystal mounting with
capillaries or fibers
n
Adjustable Z
n
Magnetic mount
n
Fits XYZ Goniometer Heads with Z Platform
or magnetic base
n
Fast, easy convenient crystal positioning
and retrieval
a p p l i c a t i o n
n
Capillary and glass fiber mounts
n
Small molecules and macromolecules
n
Available with or without a platform
a p p l i c a t i o n s
f e a t u r e s
Order Information
Cat. No. Name Description Price
HR8-028 Adjustable Crystal Mount 5 pack $58.00
A D J U S T A B L E C R Y S T A L M O U N T

3 mm brass specimen pins with platform for capillary mounting

using Four Color Mounting Clay or Red Sticky Wax. The
platform has a diameter of 9 mm which adds for extra stability in
maintaining a straight capillary for x-ray data collection. Conforms
to IUCr standards.
B R A S S S P E C I M E N P I N

d e s c r i p t i o n
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M A G N E T I C M O U N T S & S U P P O R T S
145
The Adjustable Crystal Mount is a magnetic piece (approximately
12 mm in height, 9.8 mm in diameter) designed to position a
brass specimen pin (below) and either a capillary or glass fiber
onto a goniometer head for x-ray data collection. The Adjustable
Crystal Mount is engineered to fit onto the Z Platform or magnetic
base within a goniometer head. The recessed bottom properly
positions the Crystal Mount over the center of the goniometer
head and prevents the mount from sliding across the goniometer
head during data collection. The bottom of the Crystal Mount is flat and closed to prevent the brass
specimen pin from protruding through the bottom of the Mount. The magnetic design allows for
quick, convenient, and rapid placement and retrieval of the Crystal Mount. Finally, the Crystal Mount
is designed with a single allen screw which allows the Z-translation of the brass specimen pin to
be adjusted (up and down) and then secured in position by tightening the allen screw using the
Goniometer Head Key.
Order Information
Cat. No. Name Description Price
HR4-661 Brass Specimen Pin 25 pack $32.00
HR4-663 Brass Specimen Pin with Platform 25 pack $46.00
f e a t u r e s
g o n i o m e t e r h e a d s & s u p p l i e s
Crystal of nuclear receptor.
Virginie Chavant, Structural Biology and Genomics Platform, CEBGS / IGBMC, Illkirch France.
T A B L E O F C O N T E N T S
148 x y z h e a t e d g o n i o m e t e r h e a d s
149 x y z h e a t e d g o n i o m e t e r h e a d s u p p l i e s
150 x y z s t a n d a r d g o n i o m e t e r h e a d s
151 - 152 x y z g o n i o m e t e r h e a d s u p p l i e s
153 m a g n e t i c b a s e s u p p o r t

P A G E S
n
Heated goniometer head
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Prevent icing on goniometer head & pin
n
Head height: Cryo: 34.5 mm
Capillary: 38.0 mm
n
Adjustments: X & Y: 2.5 mm
Z: 5 mm
X Y Z H E A T E D G O N I O M E T E R H E A D S
Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com 148
34.5 mm height with Z Platform magnetic base for cryocrystal-
lography. 38.0 mm height with Z Capillary for capillary mounts
(measurements from the top of a lowered Z Platform/Z Capillary
to the point where the base mount of the goniometer head
attaches to the goniometer). 2.5 mm X and Y adjustments, 5
mm Z adjustment. Ceramic heating element in base of goniometer
head maintains the head and mounting pin at a temperature that
prevents icing. Temperature controller requires 120/230 volt AC.
Head is equipped with an ACA/IUCr standard thread and support.
Each XYZ Heated Goniometer Head includes a heated goniometer head, Z capillary mount, Z Platform
9,000 gauss magnetic base, adjustment key, and storage case. Temperature controller, connection cable,
and transformer (120v USA or 230v International) are required and sold separately. Crystallization and
goniometer head accessories available separately.
Base thread is M30, 1 mm/thread.
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d e s c r i p t i o n
Cryo:
27.5 mm
XYZ Short Heated
Goniometer Head with
Cryo Mount
Capillary:
31.0 mm
XYZ Short Heated
Goniometer Head with
Capillary Mount
Heated
G
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Heated
G
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meter
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Capillary:
38.0 mm
Goniometer Head with
Capillary Mount
Cryo:
34.5 mm
Goniometer Head with
Cryo Mount
n
Heated goniometer head
n
Prevent icing on goniometer head & pin
n
Head height: Cryo: 27.5 mm
Capillary: 31.0 mm
n
Adjustments: X & Y: 2.5 mm
Z: 5 mm
27.5 mm height with Z Platform magnetic base for cryocrystal-
lography. 31.0 mm height with Z Capillary for capillary mounts
(measurements from the top of a lowered Z Platform/Z Capillary
to the point where the base mount of the goniometer head
attaches to the goniometer). 2.5 mm X and Y adjustments, 5
mm Z adjustment. Ceramic heating element in base of goniometer
head maintains the head and mounting pin at a temperature that
prevents icing. Temperature controller requires 120/230 volt AC.
Head is equipped with an ACA/IUCr standard thread and support.
Each XYZ Short Heated Goniometer Head includes a short heated goniometer head, Z capillary
mount, Z Platform 9,000 gauss magnetic base, adjustment key, and storage case. Temperature con-
troller, connection cable, and transformer (120v USA or 230v International) are required and sold
separately. Crystallization and goniometer head accessories available separately.
Base thread is M30, 1 mm/thread.
d e s c r i p t i o n
Order Information
Cat. No. Name Description Price
HR4-765 XYZ Short Heated Goniometer Head each $685.00
f e a t u r e s
f e a t u r e s
Order Information
Cat. No. Name Description Price
HR4-767 XYZ Heated Goniometer Head each $685.00
X Y Z H E A T E D G O N I O M E T E R H E A D

X Y Z S H O R T H E A T E D G O N I O M E T E R H E A D

149
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X Y Z H E A T E D G O N I O M E T E R H E A D

S U P P L I E S
Temperature Controller for the XYZ Heated and XYZ Short
Heated goniometer heads. Includes connection cable.
Transformer (power supply) for the XYZ Heated and XYZ Short
Heated goniometer heads. Available for both 120v and 230v
applications.
Replacement connection cable for the XYZ Heated and XYZ Short
Heated goniometer heads. Connects goniometer head to tempera-
ture controller.
Replacement fuse for the temperature controller.
n
Temperature controller for all XYZ heated
heads
n
Power supply for all XYZ heated heads
n
Connection cable for all XYZ heated heads
n
Fuse for temperature controllers
d e s c r i p t i o n
d e s c r i p t i o n
d e s c r i p t i o n
d e s c r i p t i o n
f e a t u r e s
f e a t u r e s
f e a t u r e s
f e a t u r e s
Order Information
Cat. No. Name Description Price
HR4-775 Temperature Controller each $98.00
Order Information
Cat. No. Name Description Price
HR4-669 24V Transformer - 120V each $41.00
HR4-755 24V Transformer - 230V each $45.00
Order Information
Cat. No. Name Description Price
HR4-657 Connection Cable each $11.00
Order Information
Cat. No. Name Description Price
HR4-777 Temperature Controller Fuse 3 pack $8.00
T E M P E R A T U R E C O N T R O L L E R F U S E
C O N N E C T I O N C A B L E
T R A N S F O R M E R
T E M P E R A T U R E C O N T R O L L E R
Cryo:
27.5 mm
XYZ Short Goniometer
Head with Cryo Mount
Capillary:
31.0 mm
XYZ Short Goniometer
Head with Capillary Mount
Capillary:
38.0 mm
Goniometer Head with
Capillary Mount
Cryo:
34.5 mm
Goniometer Head with
Cryo Mount
n
Head height: Cryo: 34.5 mm
Capillary: 38.0 mm
n
Adjustments: X & Y: 2.5 mm
Z: 5 mm
X Y Z S T A N D A R D G O N I O M E T E R H E A D S
Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com 150
34.5 mm height with Z Platform magnetic base for cryocrystal-
lography. 38.0 mm height with Z Capillary for capillary mounts
(measurements from the top of a lowered Z Platform/Z Capillary to
the point where the base mount of the goniometer head attaches
to the goniometer). 2.5 mm X and Y adjustments, 5 mm Z
adjustment. Includes XYZ Goniometer Head, Z Capillary Mount, Z
Platform 9,000 gauss magnetic base, adjustment key, and storage
case. Head is equipped with an ACA/IUCr standard thread and
support.
Base thread is M30, 1 mm/thread.
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Head height: Cryo: 27.5 mm
Capillary: 31.0 mm
n
Adjustments: X & Y: 2.5 mm
Z: 5 mm
27.5 mm height with Z Platform magnetic base for cryocrystal-
lography. 31.0 mm height with Z Capillary for capillary mounts
(measurements from the top of a lowered Z Platform/Z Capillary to
the point where the base mount of the goniometer head attaches
to the goniometer) . 2.5 mm X and Y adjustments, 5 mm Z
adjustment. Includes XYZ Short Goniometer Head, Z Capillary
mount, Z Platform 9,000 gauss magnetic base, medium (6,000
gauss) magnetic base, adjustment key, and storage case. Head is
equipped with an ACA/IUCr standard thread and support.
Base thread is M30, 1 mm/thread.
d e s c r i p t i o n
Order Information
Cat. No. Name Description Price
HR4-643 XYZ Short Standard Goniometer Head each $590.00
f e a t u r e s
f e a t u r e s
Order Information
Cat. No. Name Description Price
HR4-647 XYZ Standard Goniometer Head each $590.00
X Y Z S T A N D A R D G O N I O M E T E R H E A D

X Y Z S H O R T S T A N D A R D G O N I O M E T E R H E A D

Black plastic base threads securely into clear plastic cover and is
threaded to hold IUCr standard thread goniometer heads.
Base thread is M30, 1 mm/thread.
Goniometer head key for use with all XYZ goniometer heads.
This key is included with the purchase of any goniometer head.
Replacement set screws are available for the X and Y translation
(2 mm x 4 mm) and for Z translation collar (2 mm x 1 mm).
d e s c r i p t i o n
d e s c r i p t i o n
d e s c r i p t i o n
f e a t u r e s
f e a t u r e s
f e a t u r e s
151
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X Y Z G O N I O M E T E R H E A D S U P P L I E S
n
Plastic storage case for IUCr standard thread
goniometer heads
n
Works with all XYZ heads
n
Replacement set screws for all XYZ heads
Order Information
Cat. No. Name Description Price
HR6-100 X & Y Translation Set Screw 3 pack $9.00
HR6-102 Z Translation Collar Set Screw 3 pack $9.00
S T O R A G E C A S E F O R G O N I O M E T E R H E A D
G O N I O M E T E R H E A D K E Y
S E T S C R E W S
Order Information
Cat. No. Name Description Price
HR4-753 Storage Case for Goniometer Head each $20.00
Order Information
Cat. No. Name Description Price
HR4-659 Goniometer Head Key each $11.00
X Y Z G O N I O M E T E R H E A D S U P P L I E S
Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com 152
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Replacement Z Capillary Support for all XYZ goniometer heads.
The Adjustable Crystal Mount is a magnetic piece (approximately
12 mm in height, 9.8 mm in diameter) designed to position a brass
specimen pin (below) and either a capillary or glass fiber onto a
goniometer head for x-ray data collection. The Adjustable Crystal
Mount is engineered to fit onto the Z Platform or magnetic base
within a goniometer head. The recessed bottom properly positions
the crystal mount over the center of the goniometer head and pre-
vents the mount from sliding across the goniometer head during
data collection. The bottom of the crystal mount is flat and closed to prevent the brass specimen pin
from protruding through the bottom of the mount. The magnetic design allows for quick, convenient,
and rapid placement and retrieval of the crystal mount. Finally, the crystal mount is designed with a
single allen screw which allows the Z-translation of the brass specimen pin to be adjusted (up and
down) and then secured in position by tightening the allen screw using the Goniometer Head Key.
3 mm brass specimen pins with platform for capillary mount-
ing using Four Color Mounting Clay or Red Sticky Wax.
The platform has a diameter of 9 mm which adds extra stability in
maintaining a straight capillary for x-ray data collection. Conforms
to IUCr standards.
d e s c r i p t i o n
d e s c r i p t i o n
d e s c r i p t i o n
f e a t u r e s
n
Capillary pin support for all XYZ heads
n
Supports brass specimen pins
Order Information
Cat. No. Name Description Price
HR8-028 Adjustable Crystal Mount 5 pack $58.00
a p p l i c a t i o n
a p p l i c a t i o n s
n
Adjustable, magnetic crystal mounting with
capillaries or fibers
n
Adjustable Z
n
Magnetic mount
n
Fits XYZ Goniometer Heads with Z Platform
or magnetic base
n
Fast, easy, and convenient crystal positioning
and retrieval
n
Capillary and glass fiber mounts
n
Small molecules and macromolecules
n
Available with or without platform
f e a t u r e s
Z C A P I L L A R Y

S U P P O R T
A D J U S T A B L E C R Y S T A L M O U N T
B R A S S S P E C I M E N P I N

Order Information
Cat. No. Name Description Price
HR4-661 Brass Specimen Pin 25 pack $32.00
HR4-663 Brass Specimen Pin with Platform 25 pack $46.00
Order Information
Cat. No. Name Description Price
HR4-769 Z Capillary For all XYZ short heads - each $37.00
HR4-771 Z Capillary For all XYZ heads - each $37.00
The CrystalCap Magnetic Base (9.3 mm diameter support) is a
machined brass piece fitted with a magnet. The base fits into
the goniometer head, providing a secure base for the CrystalCap
Magnetic System. Fits most goniometer heads. The CrystalCap
Magnetic Base is available in three strengths: strong, medium, and
light. The stem of the magnet is 3 mm in diameter. The length of
the stem is 5 mm. The thickness of the platform is 3.6 mm.
Z Platform with 6,000 gauss magnet for all XYZ goniometer
heads. Less pull and reduced "snapping" than Z Platform with
9,000 gauss magnet.
Z Platform with 9,000 gauss magnet for all XYZ goniometer
heads. Same Z Platform that is included with the purchase of any
goniometer head.
d e s c r i p t i o n
d e s c r i p t i o n
d e s c r i p t i o n
f e a t u r e s
f e a t u r e s
f e a t u r e s
153
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M A G N E T I C B A S E

S U P P O R T
n
Available in 3 strengths
n
Magnetic support for CrystalCap
n
Light strength (6,000 gauss) magnetic
support for all XYZ heads
n
Magnetic support for CrystalCap
n
Strong strength (9,000 gauss) magnetic
support for all XYZ heads
n
Magnetic support for CrystalCap
Order Information
Cat. No. Name Description Price
HR4-943 Magnetic Base - Strong 9,000 gauss - each $35.00
HR4-627 Magnetic Base - Medium 6,000 gauss - each $35.00
HR4-629 Magnetic Base - Light 4,000 gauss - each $35.00
M A G N E T I C B A S E
M A G N E T I C B A S E S U P P O R T Z P L A T F O R M

- L I G H T
M A G N E T I C B A S E S U P P O R T Z P L A T F O R M

- S T R O N G
Order Information
Cat. No. Name Description Price
HR4-761 Z Platform - Strong For all XYZ short heads - each $35.00
HR4-763 Z Platform - Strong For all XYZ heads - each $35.00
Order Information
Cat. No. Name Description Price
HR4-651 Z Platform - Light For all XYZ short heads - each $37.00
HR4-653 Z Platform - Light For all XYZ heads - each $37.00
x e n o n d e r i v a t i z a t i o n
Protein Crystals.
Anja Lehwe-Litzmann, Institut for Biochemie & Biotechnologie, Martin-Luther-Universitt Halle-Wittenberg.
T A B L E O F C O N T E N T S
156 x e n o n c h a m b e r

157 x e n o n r e c o v e r y s y s t e m

P A G E S
n
Produce xenon derivatives of biological
macromolecular crystals
n
Operation pressure range of 0 to 600 psi
n
Small pressure chamber for minimum xenon
gas consumption
n
Pressure regulated safety valve
n
Safety lock prevents opening of pressurized
chamber
n
Safety shield between pressure chamber
and user
n
Quick release connectors between xenon
gas supply and pressure chamber
n
High quality xenon pressure regulator with
needle valve for fine control
n
Unique track design allows rapid crystal
transfer from xenon to dewar for freezing
n
Mini-Vial/Wick system prevents crystal
dehydration
n
CrystalCap compatible
Order Information
Cat. No. Name Description Price
HR4-791 Xenon Chamber each $2,400.00
HR4-793 Xenon Chamber Pressure Regulator each $560.00
HR4-781 Xenon Chamber Quick Female Connector each $50.00
HR4-795 Xenon Chamber Mini-Vial with Wick 5 pack $75.00
HR4-799 Xenon Chamber Vial Stand each $55.00
Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com 156
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f e a t u r e s
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Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com
The Xenon Chamber is a pressure chamber
designed to produce xenon derivatives of biologi-
cal macromolecular crystals.
Aside from the preparation of a pure sample and
subsequent crystallization, an often rate-limiting
step in the determination of biological macromo-
lecular structures by x-ray diffraction analysis is the
formation of isomorphous heavy atom derivatives.
Typically, isomorphous derivatives are formed by
diffusing heavy atoms such as lead or mercury
based compounds into the crystal with the antici-
pation that the heavy atom molecules will bind in
an ordered fashion to each biological macromol-
ecule. In order to obtain two or more successful
isomorphous derivatives, a researcher may need
to evaluate close to 100 different heavy atom
compounds by manually transferring a number of
crystals to different mother liquors each contain-
ing a different heavy atom compound.
Enter Xenon
Xenon is a noble gas which binds to specific sites
in a biological macromolecule. There are numer-
ous examples demonstrating that xenon-macro-
molecule complexes can serve as heavy atom
derivatives.
1-8
The xenon-macromolecule complex
is obtained by placing the macromolecular crystal
under a pressurized xenon atmosphere. A clear
advantage in using xenon is that one can simply
screen xenon by pressurizing a loop-mounted
crystal rather than having to set up plates and hav-
ing to transfer crystals from numerous drops.
Enter the Xenon Chamber
The Xenon Chamber is a simple, yet effective
device designed to pressurize loop-mounted bio-
logical macromolecular crystals in the presence of
xenon gas at room temperature. Crystals mounted
in loops such as the CrystalCap system are placed
into the Xenon Chamber. Once sealed, the cham-
ber is pressurized with xenon gas so that the
crystal and macromolecules are equilibrated in
a vapor saturated xenon atmosphere. Following
depressurization of the chamber, the loop-mount-
ed crystal is simply lifted and slid along the Xenon
Chamber track and quickly lowered into a dewar
for freezing in liquid nitrogen or propane. The
transfer from pressurized xenon to the freeze is
accomplished in seconds.
Specifications
The Xenon Chamber measures 14 1/4" tall, 13 1/2"
wide, and 10 1/4" deep. The pressure chamber
measures 9.5 mm (diameter) by 29 mm (depth).
The Xenon Chamber has an operation pressure
range of 0 to 600 psi (0 to 4.1 Mpa, 0 to 41 Bar).
The pressure regulator for the Xenon Chamber
is designed to be used with inert gases such
as xenon. The CGA number for the pressure
regulator is 580. The pressure regulator has a hose
length of 21" with a quick connector on the end to
attach it to the Xenon Chamber.
Each Xenon Chamber comes complete with a
CrystalCap Dewar Stand that fits the 1,000 ml
Dewar Flask (HR4-699) and a pack of five Mini-
Vials with Wicks. The Xenon Chamber Pressure
Regulator is sold separately. Additional Mini-Vials
with Wick and Vial Stands may also be obtained
separately.
References
1. PNAS, Oct. 12 1999, Vol.96, No. 21, 11717-11722.
2. Soltis, S.M., Stowell, M.H.B., Wiener, M.C., Phillips, G.N. Jr., & Rees, D.C.,
J. Appl. Cryst. (1997) 30, 190-194.
3. Stowell, M.H.B., Soltis, S.M., Kisker, C., Peters, J.W., Schindelin, H., Rees,
D.C., Cascio, D., Beamer, L., Hart, P.J., Wiener, M.C., & Whitby, F.G., J.
Appl. Cryst. (1996) 29, 608-613.
4. Schiltz, M., Fourme, R., Broutin, I., & Prange, T., Structure (1995) 3,
309-316.
5. Schitz, M., Prange, T., & Fourme, R., J. Appl. Cryst. (1994) 27, 950-960.
6. Vitali, J., Robbins, A.H., Almo, S.C., & Tilton, R.F., J. Appl. Cryst. (1991)
24, 931-935.
7. Schiltz, M., Shepard, W., Fourme, R., Prange, T., De La Fortelle, R., &
Bricogne, G., Acta Cryst. (1997) D53, 78-92.
8. Otwinowski, Z. & Minor, W., Methods Enzymol. Vol. 276, edited by C.W.
Carter, Jr. and R.M. Sweet. New York: Academic Press (1997).
d e s c r i p t i o n
X E N O N C H A M B E R

C A S E S T U D Y : X E N O N D E R I V A T I V E
& C R Y S T A L S T R U C T U R E
157
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A Xenon Derivative of Tt UvrB Crystal Prepared using the
Hampton Research Xenon Chamber
d e s c r i p t i o n
Thermus thermophilus UvrB (Tt UvrB), a 76 kDa protein is a com-
ponent of the nucleotide excision repair system. The Tt UvrB crystal
structures was determined by single isomorphous replacement /
anomalous scattering from a xenon derivative. The UvrB crystals
were pressurized with 500 psi of xenon gas for 15 minutes at room
temperature and flash cooled for data collection. Xenon derivatized
crystals diffracted to 1.9 angstrom.
Reference:
Machius, M., Henry, L., Palnitkar, M., and Deisenhofer, J. (1999) Proc. Natl. Acad. Sci USA, 96,
11717-11722.
Overall structure
of Tt UvrB with
the Xenon sites
(red spheres).
Environment of
Xenon Site 1
with Fo-Fc densi-
ty for the Xenon
atom contoured
at 10 .
Environment of
Xenon Site 2
with Fo-Fc densi-
ty for the Xenon
atom contoured
at 10 .
Environment of
Xenon Site 3
with Fo-Fc densi-
ty for the Xenon
atom contoured
at 10 .
Environment of
Xenon Site 4
with Fo-Fc densi-
ty for the Xenon
atom contoured
at 10 .
When using the Xenon Chamber to obtain deriva-
tized crystals, the gas pressure is typically between
200 and 500 psi. A typical 50 liter compressed
xenon gas cylinder has an initial pressure of 600
psi. Without some type of recovery system, if the
cylinder pressure is less than desired, the cylinder
cannot be used and the expensive xenon gas is
wasted. With the Xenon Recovery System connected
to a compressed xenon gas cylinder, pressures can
be increased by a factor of 9. This means a typical
compressed xenon gas cylinder is utilized at 95%
efficiency versus 50% efficiency without the recovery
system. The system easily connects to the Xenon
Chamber and Pressure Regulator. Contact Technical
Support for complete specifications.
Order Information
Cat. No. Name Description Price
HR4-797 Xenon Recovery System each $1,200.00
d e s c r i p t i o n
a p p l i c a t i o n
f e a t u r e s
n
Scavenge and recover xenon gas from low
pressure xenon gas cylinders
n
Utilize up to 95% of xenon gas in cylinders
X E N O N R E C O V E R Y S Y S T E M

l a b e l s & p e n s
Proteins crystals.
Kasumi Kobayashi, Taiji Nakae and Hiroyuki Akama. The Kitasato Institute, Kanagawa, Japan.
T A B L E O F C O N T E N T S
160 p e ns
160 - 161 t o u g h - t a g s

o n a r o l l
161 c r y o - b a b i e s

a n d c r y o - t a g s

o n a r o l l
162 t o u g h - s p o t s

o n a r o l l
163 l a s e r c r y o - b a b i e s

a n d c r y o - t a g s

163

l a s e r t o u g h - s p o t s

P A G E S
Designed for use with 0.2 ml microtubes, Tough-Tags On a Roll
- Teeny will withstand thermal-cycling, autoclaving, boiling water
baths and freezing to -40C.
Label Size: 0.81" x 0.28" (21 mm x 7 mm)
Color: White; Quantity: 1,500 labels per roll
d e s c r i p t i o n
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O N A R O L L - T E E N Y
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Labels for 0.2 ml microtubes
a p p l i c a t i o n
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Marking crystallization plates, labels,
sample tubes, and vials
n
Marking of labels and samples
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Fine point writing tip
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Black ink
n
Fast drying, permanent ink
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Red ink or black ink
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Pre-cut, peel off labels
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Withstands solvents, incubation in high
humidity, and freezing to -40C
n
Works best with extra fine point markers
(Teeny Writer)
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Convenient, easy to dispense roll
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T O U G H - T A G S

O N A R O L L
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HR4-458 Tough-Tags

On a Roll - Teeny 1 roll $35.00

Order Information
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HR4-462 Teeny Writer each $6.00
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HR4-464 Solvent Resistant Pen - Black each $6.00
HR4-466 Solvent Resistant Pen - Red each $6.00
T E E N Y W R I T E R
S O L V E N T R E S I S T A N T P E N
The Teeny Writer has an extra fine tip. Recommended for writing on
all our labels for samples that will be stored at room temperature and
not exposed to chemicals and/or caustic agents. Ink dries in a matter
of seconds. The ink is NOT resistant to detergents and reagents used
in histology procedures. Color: Black.
The Solvent Resistant Pen is recommended for writing on Tough-
Tags. The ultra-permanent ink dries in a matter of seconds and
resists solvents such as xylene, alcohol, acetone, and formalin.
Testing is advised before use.
d e s c r i p t i o n
d e s c r i p t i o n
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T O U G H - T A G S

O N A R O L L
161
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HR4-460 Tough-Tags

On a Roll - Sidewall 1 roll $35.00

Tough-Tags On a Roll - Sidewall have been carefully engineered to
strongly adhere to all plastics and other materials. Sidewall Tough-
Tags withstand organic solvents, caustic agents, humid incubators,
boiling water baths, and freezer temperatures without peeling.
These pre-cut labels fit perfectly onto the sides of crystallization
plates allowing for visual identification of the individual plates in
a stack.
Label Size: 1.50" x 0.25" (38 mm x 6 mm)
Color: White; Quantity: 1,000 labels per roll
n
Labels for crystallization plates
a p p l i c a t i o n
T O U G H - T A G S

O N A R O L L S I D E WA L L
d e s c r i p t i o n
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Withstands solvents, incubation in high
humidity, and freezing to -20C
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Easy to peel off and affix
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Pre-cut labels fit perfectly on the side of
crystallization plates
f e a t u r e s
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Freezable in liquid nitrogen
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Chemically resistant
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Made of a special smear-resistant material
that will accept any marking instrument
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Pre-cut labels on a roll in dispenser box
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HR4-404 Cryo-Babies

On a Roll 1 roll $36.00

HR4-406 Cryo-Tags

On a Roll 1 roll $36.00

Labels for cryogenic storage. Withstand conventional and cryo-
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The labels will also withstand boiling water baths to 100C and dry
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metals. They resist detergents, oils, solvents, caustic agents, and
other challenges without peeling or falling off. Great for labeling
plastics and other materials for cryogenic storage. Cryo-Babies are
ideal for1.5-2.0 ml or larger tubes. Apply label to tube or container
BEFORE freezing.
Catalog #HR4-404 Label Size: 1.28" x 0.50" (33 mm x 13 mm)
Catalog #HR4-406 Label Size: 1.50" x 0.75" (38 mm x 19 mm)
Color: White; Quantity: 1,000 labels per roll
n
Labels for cryo labware cryogenic storage
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C R Y O - B A B I E S

A N D C R Y O - T A G S

O N A R O L L
T O U G H - S P O T S

O N A R O L L
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HR4-436 Tough-Spots

On a Roll - Small White - 1 roll $31.00

HR4-438 Tough-Spots

On a Roll - Small Red - 1 roll $31.00

HR4-440 Tough-Spots

On a Roll - Small Yellow - 1 roll $31.00

HR4-442 Tough-Spots

On a Roll - Small Green - 1 roll $31.00

HR4-444 Tough-Spots

On a Roll - Small Blue - 1 roll $31.00

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HR4-446 Tough-Spots

On a Roll - Large White - 1 roll $31.00

Tough-Spots On a Roll - Small are chemically inert, polyvinyl labels
that adhere to all plastics. They are 3/8" diameter circular labels
that are useful for tagging and easily identifying a wide range of
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colors.
Label Size: 3/8" diameter (9.5 mm diameter)
Quantity: 1,000 labels per roll
Tough-Spots On a Roll - Large are chemically inert, polyvinyl
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Label Size: 1/2" diameter (13 mm diameter)
Color: White; Quantity: 1,000 labels per roll
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O N A R O L L - S M A L L
T O U G H - S P O T S

O N A R O L L - L A R G E
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a p p l i c a t i o n
a p p l i c a t i o n
d e s c r i p t i o n
d e s c r i p t i o n
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For microtube cap or lid identification
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Freezable in liquid nitrogen
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Autoclavable and boilable
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Chemically resistant
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Won't dry out, fall off, or tear
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For microtube cap or lid identification
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Freezable in liquid nitrogen
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Won't dry out, fall off, or tear
f e a t u r e s
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L A S E R C R Y O - B A B I E S

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163
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1 pack $70.00
Labels for cryogenic storage. Withstand conventional and cryogenic
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labels will also withstand boiling water baths to 100C and dry heat
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als. They resist detergents, oils, solvents, caustic agents, and other
challenges without peeling or falling off. Ideal for 1.5 to 2.0 ml or
larger tubes.
Label Size: 1.28 x 0.50" (33 mm x 13 mm)
Color: White; Quantity: 85 labels per sheet (1,700/pack)
Labels for cryogenic storage. Withstand conventional and cryogenic
storage (vapor and liquid phase nitrogen storage) to -196C. The
labels will also withstand boiling water baths to 100C and dry
heat up to 150C. Polyolefin labels adhere to all plastics, glass, and
metals. They resist detergents, oils, solvents, caustic agents, and
other challenges without peeling or falling off. Ideal for labeling
plastics and other materials for cryogenic storage.
Label Size: 1.50 x 0.75" (38 mm x 19 mm)
Color: White; Quantity: 60 labels per sheet (1,200/pack)
Tough-Spots Small are chemically inert, polyvinyl labels that adhere
to all plastics. They are 3/8" diameter circular labels that are useful
for tagging and easily identifying a wide range of small containers in
the crystallization lab.
Label Size: 3/8" diameter (9.5 mm diameter)
Color: White; Quantity: 192 labels per sheet (3,840/pack)
n
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f e a t u r e s
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L A S E R T O U G H - S P O T S

Here is a recipe to try:

Mosaicity is about 0.5
t0 0.6
Reagent:
Crystal Screen Cryo
Reagent 23
Mix equal amounts of
Glucose Isomerase
and reagent. Vapor
diffusion method.

Mount crystal in
CryoLoop.

Mosaicity may be
a bit more in this
reagent and the
unit cell will shrink a
b o o k s
Proteins crystals.
Kasumi Kobayashi, Taiji Nakae and Hiroyuki Akama. The Kitasato Institute, Kanagawa, Japan.
T A B L E O F C O N T E N T S
166 m a c r o m o l e c u l a r c r y s t a l l o g r a p h y
166 m a c r o m o l e c u l a r c r y s t a l l o g r a p h y p r o t o c o l s ,
v o l u m e 1
167 p r o t e i n c r y s t a l l i z a t i o n s t r a t e g i e s f o r
s t r u c t u r a l g e n o m i c s
167 p r a c t i c a l p r o t e i n c r y s t a l l o g r a p h y
167 p r e p a r a t i o n a n d a n a l y s i s o f p r o t e i n c r y s t a l s
168 p r o t e i n m e t h o d s
168 m e t h o d s i n e n z y m o l o g y - p a r t c
168 m e t h o d s i n e n z y m o l o g y - p a r t d
169 c r y s t a l l o g r a p h y m a d e c r y s t a l c l e a r
169 o u t l i n e o f c r y s t a l l o g r a p h y f o r b i o l o g i s t s
169 i n t r o d u c t i o n t o m a c r o m o l e c u l a r c r y s t a l l o g r a p h y
170 p r i n c i p l e s o f p r o t e i n x - r a y c r y s t a l l o g r a p h y
170 m e m b r a n e p r o t e i n p u r i f i c a t i o n &
c r y s t a l l i z a t i o n , a p r a c t i c a l g u i d e
170 m e t h o d s a n d r e s u l t s i n c r y s t a l l i z a t i o n o f
m e m b r a n e p r o t e i n s
171 m a c r o m o l e c u l a r c r y s t a l l i z a t i o n m e t h o d s ,
v o l u m e 3 4 , n u m b e r 3
171 j o u r n a l o f s t r u c t u r a l b i o l o g y
172 p r e s e n t a t t h e f l o o d :
h o w s t r u c t u r a l m o l e c u l a r b i o l o g y c a m e a b o u t
172 c r y s t a l s a n d l i f e : a p e r s o n a l j o u r n e y
172 p r o t e i n c r y s t a l l o g r a p h y : a c o n c i s e g u i d e
173 p r o t e i n c r y s t a l l i z a t i o n
173 v i e w e r s
P A G E S
B O O K S
Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com 166 Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com
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Covers all aspects of protein crystallography. The 386 page text
is a practical handbook intended to be used by anyone who
wants to solve a structure by protein crystallography. This book
is divided into four chapters: (1) Laboratory Techniques, (2) Data
Collection Techniques, (3) Computational Techniques, and (4)
Protein Crystallography Cookbook.
P R A C T I C A L P R O T E I N C R Y S T A L L O G R A P H Y
n
Authors: Mark Sanderson and Jane Skelly
n
Publisher: Oxford University Press, 2007
n
ISBN-10: 0198520972
n
ISBN-13: 978-0198520979
n
Hardcover: 400 pages
d e s c r i p t i o n
Macromolecular Crystallography - conventional and high-through-
put methods
Macromolecular crystallography is the study of macromolecules
(proteins and nucleic acids) using x-ray crystallographic techniques
in order to determine their molecular structure. The knowledge of
accurate molecular structures is a pre-requisite for rational drug
design, and for structure-based function studies to aid the devel-
opment of effective therapeutic agents and drugs. The successful
determination of the complete genome (genetic sequence) of several species (including humans) has
recently directed scientific attention towards identifying the structure and function of the complete
complement of proteins that make up that species; a new and rapidly growing field of study called
'structural genomics'. There are now several important and well-funded global initiatives in operation
to identify all of the proteins of key model species. One of the main requirements for these initiatives
is a high-throughput crystallization facility to speed-up the protein identification process. The extent to
which these technologies have advanced, calls for an updated review of current crystallographic theory
and practice. This practical reference book features the latest conventional and high-throughput meth-
ods, and includes contributions from a team of internationally recognized leaders and experts. It will
be of relevance and use to graduate students, research scientists and professionals currently working
in the field of conventional and high-throughput macromolecular crystallography.
Author Information
Mark Sanderson is an experienced protein and nucleic-acid crystallographer and co-editor of Methods
in Molecular Biology, who has solved several key ab initio structures, notably nucleic-acid proteins.
Jane Skelly has widespread experience in the expression and crystallisation of macromolecules. Both
authors lecture undergraduates in structural biology.
M A C R O M O L E C U L A R C R Y S T A L L O G R A P H Y
d e s c r i p t i o n
Order Information
Cat. No. Name Description Price
HR5-224 Macromolecular Crystallography each $156.00
n
Author: Sylvie Doublie
n
Publisher: Humana Press, 2007
n
ISBN-10: 1588292924
n
ISBN-13: 978-1588292926
n
Hardcover: 312 pages
d e s c r i p t i o n
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Cat. No. Name Description Price
HR5-215 Macromolecular Crystallography Protocols, Vol. 1 each $40.00
Macromolecular Crystallography Protocols, now in two volumes,
examines major developments that have occurred since publica-
tion of the acclaimed first edition nearly a decade ago. Volume 1,
Preparation and Crystallization of Macromolecules and Volume 2,
Structure Determination, explore recent advances that have acceler-
ated the pace of structural determination and made crystallography
accessible to a broader range of investigators. Volume 1 is composed
of detailed protocols for the preparation and optimization of crystals,
including tips from the experts on the best methods for inducing proteins to adopt their crystalline form.
Volume 2 complements the first volume by addressing laboratory techniques for crystal handling and
structural characterization, as well as computational techniques for data collection, phasing, and refinement.
The volume concludes with a detailed and insightful survey of available crystallographic software. These
volumes will be an indispensable reference for obtaining macromolecular crystals and determining their
three-dimensional structure.
M A C R O M O L E C U L A R C R Y S T A L L O G R A P H Y P R O T O C O L S , V O L U M E 1
167
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Research advances in recent years have opened up the scope for the
development of new methods and tools to overcome the bottleneck
of protein crystallization. A variety of parameters that could previously
not be explored are now accessible thanks to sophisticated apparatus
and the development of new science-based techniques to monitor and
control the process of crystallization. However, in order to become
useful to the structural genomics effort, it is vital to miniaturize and
automate these techniques and adapt them to cope with the vast
numbers of leads resulting from the high-throughput screening procedures. Such efforts are those of the
immediate future and the focus of this book.
P R O T E I N C R Y S T A L L I Z A T I O N S T R A T E G I E S F O R S T R U C T U R A L G E N O M I C S
n
Editor: Naomi E. Chayen
n
Publisher: International University Line,
2007
n
ISBN-10: 097207743X
n
ISBN-13: 978-0972077439
n
Hardcover: 290 pages
d e s c r i p t i o n
P R O T E I N C R Y S T A L L I Z A T I O N T E C H N I Q U E S , S T R A T E G I E S , & T I P S
Order Information
Cat. No. Name Description Price
HR5-230 Protein Crystallization Strategies for Structural Genomics each $109.00
Covers all aspects of protein crystallography. The 386 page text
is a practical handbook intended to be used by anyone who
wants to solve a structure by protein crystallography. This book
is divided into four chapters: (1) Laboratory Techniques, (2) Data
Collection Techniques, (3) Computational Techniques, and (4)
Protein Crystallography Cookbook.
P R A C T I C A L P R O T E I N C R Y S T A L L O G R A P H Y
n
Author: Duncan E. McRee
n
Publisher: Academic Press; 2nd edition, 1999
n
ISBN-10: 0124860524
n
ISBN-13: 978-0124860520
n
Hardcover: 504 pages
d e s c r i p t i o n
Covers all aspects of protein crystallography. It is a practical
handbook intended to be used by anyone who wants to solve
a structure by protein crystallography. This book is divided into
four chapters: (1) Laboratory Techniques; (2) Data Collection
Techniques; (3) Computational Techniques; and (4) Protein
Crystallography Cookbook.
P R A C T I C A L P R O T E I N C R Y S T A L L O G R A P H Y
d e s c r i p t i o n
Order Information
Cat. No. Name Description Price
HR5-211 Practical Protein Crystallography each $142.00
This hard-to-find book is a classic. The first text written spe-
cifically for macromolecular crystallization. This twelve chapter
book provides an interface between the techniques and prac-
tices common to most biochemists and the procedures familiar
to x-ray diffractionists.
n
Author: Alexander McPherson
n
Publisher: Krieger Publishing, 1989
n
ISBN-10: 089464355X
n
ISBN-13: 978-0894643552
n
Hardcover: 384 pages
d e s c r i p t i o n
P R E P A R A T I O N A N D A N A L Y S I S O F P R O T E I N C R Y S T A L S
Order Information
Cat. No. Name Description Price
HR5-213 Preparation and Analysis of Protein Crystals each $75.00
B O O K S
Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com 168 Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com
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P R O T E I N M E T H O D S
n
Authors: Daniel M. Bollag, Michael D.
Rozycki, & Stuart J. Edelstein
n
Publisher: Wiley-Liss Publishing; 2nd edition,
1996
n
ISBN-10: 0471118370
n
ISBN-13: 978-0471118374
n
Paperback: 415 pages
d e s c r i p t i o n
Order Information
Cat. No. Name Description Price
HR5-223 Protein Methods each $98.00
This text is a handy and practical guide of protein methods which
includes 36 pages dedicated to a straightforward, practical introduction
to macromolecular crystallization. Chapter titles include: Preparation
for Protein Isolation, Protein Extraction and Solubilization, Protein
Concentration Determination, Concentrating Protein Solutions, Gel
Electrophoresis, IEF, Immunoblotting, Ion Exchange, Affinity and Gel
Filtration Chromatography, and Hanging Drop Crystallization.
Macromolecular Crystallography, Part C (Methods in Enzymology,
Volume 368). Topics covered include: methodological methods in
crystals and data analysis.
M E T H O D S I N E N Z Y M O L O G Y - P A R T C
n
Editor: Charles W. Carter, Jr.
n
Publisher: Elsevier, 2003
n
ISBN-10: 0121822710
n
ISBN-13: 978-0121822712
n
Hardcover: 368 pages
d e s c r i p t i o n
Order Information
Cat. No. Name Description Price
HR5-210 Methods in Enzymology - Vol 368, Part C each $150.00
Macromolecular Crystallography, Part D (Methods in Enzymology,
Volume 374). Topics covered include: Phases, Map Interpretation
and Refinement, Analysis and Software.
M E T H O D S I N E N Z Y M O L O G Y - P A R T D
n
Editor: Charles W. Carter, Jr.
n
Publisher: Elsevier, 2003
n
ISBN-10: 0121827771
n
ISBN-13: 978-0121827779
n
Hardcover: 679 pages
d e s c r i p t i o n
Order Information
Cat. No. Name Description Price
HR5-212 Methods in Enzymology Volume 374, Part D each $150.00
169
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This book is an exciting resource for those who are vitally inter-
ested in macromolecular models derived from x-ray crystallography
but are unfamiliar with, or daunted by, crystallography itself. This
book is for readers who want an intellectually satisfying under-
standing of how three-dimensional models of proteins and nucleic
acids are derived from x-ray diffraction analysis of crystals. This
understanding is essential for wise use of crystallographic models
in studying enzyme catalysis, protein-mediated transport and
recognition, protein/nucleic acid interaction, and protein folding.
It is also vital in interpreting data from chemical, kinetic, thermodynamic, and spectroscopic studies
of macromolecules.
C R Y S T A L L O G R A P H Y M A D E C R Y S T A L C L E A R
n
Author: Gale Rhodes
n
Publisher: Academic Press; 3rd edition, 2006
n
ISBN-10: 0125870736
n
ISBN-13: 978-0125870733
n
Paperback: 352 pages
d e s c r i p t i o n
Order Information
Cat. No. Name Description Price
HR5-233 Crystallography Made Crystal Clear each $63.00
P R A C T I C A L P R O T E I N C R Y S T A L L O G R A P H Y
d e s c r i p t i o n
Outline of Crystallography for Biologists is intended for researchers
and students in the biological sciences who require an insight into
the methods of x-ray crystallography without needing to learn all the
relevant theory. The main text is purely descriptive and is readable
by those with minimal mathematical knowledge. Some mathematical
detail is given throughout in boxes, but these can be ignored. Theory
is limited to the essentials required to comprehend issues of quality.
There is an extensive reference section and suggestions for further
reading for those who wish to delve deeper. The first part, 'Fundamentals', presents the underlying ideas
which allow x-ray structure analysis to be carried out and provides an appropriate background to courses
in structural determination. The second part, 'Practice', gives more information about the procedures
employed in the course of crystal structure determination. The emphasis is on the quality measures of
x-ray diffraction analysis to give the reader a critical insight into the quality and accuracy of a structure
determination and to enable the reader to appreciate which parts of a structure determination may have
caused special difficulty. There is no pretence of completeness and many matters discussed in standard
crystallography texts are deliberately omitted. However, issues not brought out in the standard texts are
discussed, making it a useful resource for non-practicing crystallographers as well.
n
Author: David Blow
n
Publisher: Oxford University Press, 2002
n
ISBN-10: 0198510519
n
ISBN-13: 978-0198510512
n
Paperback: 248 pages
Order Information
Cat. No. Name Description Price
HR8-086 Outline of Crystallography for Biologists each $82.00
O U T L I N E O F C R Y S T A L L O G R A P H Y F O R B I O L O G I S T S
n
Author: Alexander McPherson
n
Publisher: Wiley-Blackwell; 2nd edition,
2009
n
ISBN-10: 0470185902
n
ISBN-13: 978-0470185902
n
Paperback: 267 pages
d e s c r i p t i o n
Order Information
Cat. No. Name Description Price
HR8-062 Introduction to Macromolecular Crystallography each $80.00
This book provides a comprehensive, approachable summary of the
field of crystallography, from the fundamental theory of diffraction
and properties of crystals to applications in determining macromo-
lecular structure.
Dedicated to providing a complete introduction to the subject
that does not assume a background in physics or math, the book's
contents flow logically from basic principles to key methods, such as
those for solving phase problems, interpretation of Patterson maps,
and the difference Fourier method. The Introduction includes:
Practical Instruction on the Interpretation of Data and Methods for Determining Phases.
I N T R O D U C T I O N T O M A C R O M O L E C U L A R C R Y S T A L L O G R A P H Y
Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com 170 Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com
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B O O K S
M E M B R A N E P R O T E I N P U R I F I C AT I O N & C R Y S TA L L I Z AT I O N , A P R A C T I C A L G U I D E
Order Information
Cat. No. Name Description Price
HR5-241 Membrane Protein Purification and Crystallization each $98.00
This guide presents isolation and crystallization techniques in a
concise form, emphasizing the critical aspects unique to membrane
proteins. It explains the principles of the methods and provides
protocols of general use, permitting researchers and students new
to this area to adapt these techniques to their particular needs. Key
features: Provides general guidelines and strategies for isolation
and crystallization of membrane proteins. Gives detailed protocols
that have wide application, and low specialized equipment needs.
Emphasizes recent progress in production and purification of recombinant membrane proteins, espe-
cially of histidine-tagged and other affinity-epitope-tagged proteins. Summarizes recent developments of
Blue-Native PAGE, a high resolution separation technique, which is independent of the use of recombi-
nant techniques, and is especially suited for proteomic analyses of membrane protein complexes. Gives
detailed protocols for membrane protein crystallization, and describes the production and use of anti-
body fragments for high resolution crystallization. Presents a comprehensive guide to 2D-crystallization
of membrane proteins.
n
Editors: Carola Hunte, Gebhard Von
Jagow, Hermann Schagger
n
Publisher: Academic Press;
2nd edition, 2002
n
ISBN-10: 0123617766
n
ISBN-13: 978-0123617767
n
Spiral-bound: 316 pages
d e s c r i p t i o n
P R I N C I P L E S O F P R O T E I N X - R A Y C R Y S T A L L O G R A P H Y
n
Author: Jan Drenth
n
Publisher: Springer; 3rd edition, 2006
n
ISBN-10: 0387333347
n
ISBN-13: 978-0387333342
n
Hardcover: 332 pages
d e s c r i p t i o n
Order Information
Cat. No. Name Description Price
HR5-219 Principles of Protein X-Ray Crystallography each $90.00
Strong on crystallography, light on crystal growing (17 pages).
Fifteen chapters including Crystallizing a Protein, X-Ray Sources
and Detectors, Crystals, Theory of X-Ray Diffraction By a Crystal,
Average Reflection Intensity and Distribution of Structure Factor
Data, Special Forms of the Structure Factor, Phase Improvement,
The Solution of the Phase Problem by the Isomorphous Replacement
Method, Anamolous Scattering, Molecular Replacement, Direct
Methods, Refinement, Combination of Phase Information, Phase
Information from Partial Structure Data, Solvent Flattening, and
Molecular Averaging.
M E T H O D S A N D R E S U LT S I N C R Y S T A L L I Z AT I O N O F M E M B R A N E P R O T E I N S
n
Editor: So Iwata
n
Publisher: International University Line,
2002
n
ISBN-10: 0963681796
n
ISBN-13: 978-0963681799
n
Hardcover: 380 pages
d e s c r i p t i o n
This text consists of four parts. Parts 1 and 2 include an introduc-
tion as well as general principles and techniques in membrane pro-
tein crystallization. This is followed by chapters covering the use of
detergents, crystallization in lipidic cubic phases, the use and gen-
eration of antibody fragments, porin as a model, and crystallization
of membrane proteins in oils. Part 3 focuses on specific examples
of membrane proteins whose structures have been solved. Part 4
covers the design of a kit for membrane protein crystallization.
Order Information
Cat. No. Name Description Price
HR5-237 Methods and Results in Crystallization of Membrane Proteins each $109.00
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This is a single issue of Methods, a companion journal to Methods
in Enzymology, focusing on rapidly developing techniques. This
single issue is dedicated to Macromolecular Crystallization and be-
gins with an Introduction to Protein Crystallization by Alexander
McPherson. The issue features the following additional thirteen
articles. Protein crystallization and phase diagrams, Neer Asherie.
Growth and disorder of macromolecular crystals: insights from
atomic force microscopy and x-ray diffraction studies, Alexander J.
Malkin and Robert E. Thorne. Ions from the Hofmeister series and
osmolytes: effects on proteins in solution and in the crystallization process, Kim D. Collins. Effects of
naturally occurring osmolytes on protein stability and solubility; issues important in protein crystalliza-
tion, D.W. Bolen. Practical aspects of using the microbatch method in screening conditions for protein
crystallization, Allan DArcy, Aengus Mac Sweeney, and Alexander Haber. Automated systems for protein
crystallization, Joel Bard, Kimberly Ercolani, Kristine Svenson, Andrea Olland, and Will Somers. Lipidic
cubic phases as matrices for membrane protein crystallization, Peter Nollert. The use of recombinant
and molecular engineering in protein crystallization, Zygmunt Derewenda. A pedestrian guide to mem-
brane protein crystallization, Michael Wiener. Crystallization data mining in structural genomics: using
positive and negative results to optimize protein crystallization screens, Rebecca Page and Raymond C.
Stevens. Predictive models for protein crystallization, Bernhard Rupp and Junwen Wang. Crystallization
of RNA and RNA-protein complexes, Ailong Ke and Jennifer A. Doudna. Macromolecular cryocrystallog-
raphy methods for cooling and mounting protein crystals are cryogenic temperatures, J.W. Pfugrath.
n
Edited by: Alexander McPherson
n
Publisher: Elsevier; Methods, Volume 34,
Number 3, November 2004
n
ISSN: 1046-2023
n
Paperback: 172 pages
Order Information
Cat. No. Name Description Price
HR8-061 Macromolecular Crystallization each $40.00
d e s c r i p t i o n
M A C R O M O L E C U L A R C R Y S T A L L I Z A T I O N M E T H O D S , V O L U M E 3 4 , N U M B E R 3
This special issue of the Journal of Structural Biology focuses on
Macromolecular Crystallization in the Structural Genomics Era.
Following an Introduction by Alexander McPherson, there are 19
regular articles focused on the crystallization of biological macro-
molecules. If crystallization is your thing, you need this. Thanks to
everyone contributing to this special issue. Well done everyone!
J O U R N A L O F S T R U C T U R A L B I O L O G Y
n
Editor: Alexander McPherson
n
Publisher: Academic Press; Volume 142,
Number 1, April 2003
n
ISSN: 1047-8477
n
Paperback: 231 pages
d e s c r i p t i o n
Order Information
Cat. No. Name Description Price
HR8-100 Journal of Structural Biology each $60.00
Crystal image.

Sylvia Luckner, Lehrstuhl fr Biotechnik,
University Erlangen-Nrnberg Germany.
B O O K S
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Order Information
Cat. No. Name Description Price
HR5-214 Present at the Flood each $35.00
This book chronicles a revolution in molecular biologythe crucial
30 years (roughly between 1933 and 1963) during which our ideas
about proteins and nucleic acids changed from those of formless,
functionless organic chemicals into precisely structured molecular
machines with specifc biological purpose. Proteins evolved from
being colloidal micelles or globules with no specifc structure (or
even sequence) into quite precisely structured molecular catalysts,
carrier proteins, and information-sensing agents. Indeed, the very
idea that the amino acids of a protein were linked in a specifc
order in long linear chains was not accepted initially. During this same time period, DNA changed
from being a sterile repeating polymer of no particular function (the tetranucleotide hypothesis) into a
double helix that serves as the archive of genetic information.
n
Author: Richard Dickerson
n
Publisher: Sinauer Associates, 2005
n
ISBN-10: 0878931686
n
ISBN-13: 978-0878931682
n
Paperback: 307 pages
d e s c r i p t i o n
PRESENT AT T HE F L OOD: HOW ST RUCT URAL MOL ECUL AR BI OL OGY CAME ABOUT
n
Author: Cele Abad-Zapatero
n
Publisher: International University Line,
2002
n
ISBN-10: 0972077405
n
ISBN-13: 978-0972077408
n
Paperback: 300 pages
d e s c r i p t i o n
Order Information
Cat. No. Name Description Price
HR5-239 Crystals and Life: A Personal Journey each $15.00
This book introduces crystallography, the field, and applications
of structural biology using metaphors from the arts, music,
poetry, and architecture. The reader will find a tapestry filled
by personal experiences, historical anecdotes, biographical
snapshots of scientific heroes of the field, and crystallographic
concepts illustrated with artistic analogies. All these elements
create a concoction that makes crystallography accessible,
comprehensible, intriguing, inspiring, and beautiful. The book
is divided in sections covering the following: the basic elements
of crystallography, novel technologies, practical applications and future perspectives. Since crystal-
lography is quintessentially a visual science, the illustrations play a very important role in providing
an excellent set of landmarks to guide the reader on this personal journey of discovery.
C R Y S T A L S A N D L I F E : A P E R S O N A L J O U R N E Y
n
Authors: Eaton E. Lattman and
Patrick J. Loll
n
Publisher: The Johns Hopkins University
Press, 2008
n
ISBN-10: 0801888085
n
ISBN-13: 978-0801888085
n
Paperback: 152 pages
d e s c r i p t i o n
Order Information
Cat. No. Name Description Price
HR5-221 Protein Crystallography: A Concise Guide each $37.00
The proteome remains a mysterious realm. Researchers have
determined the structures of only a small fraction of the proteins
encoded by the human genome. Crystallography continues to
be the primary method used to determine the structures of the
remaining unknown proteins. This imaging technique uses the
diffraction of x-rays to determine a proteins three-dimensional
molecular structure. Drawing on years of research and teach-
ing experience, Eaton E. Lattman and Patrick J. Loll use clear
examples and abundant illustrations to provide a concise and accessible primer on protein crystal-
lography. Discussing the basics of diffraction, the behavior of two- and three-dimensional crystals,
phase determination (including MIR and MAD phasing and molecular replacement), the Patterson
function, and refinement, Lattman and Loll provide a complete overview of this important tech-
nique, illuminated by physical insights. The crisp writing style and simple illustrations will provide
beginner crystallographers with a guide to the process of unraveling protein structure.
P R O T E I N C R Y S T A L L O G R A P H Y : A C O N C I S E G U I D E
Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com
Order Information
Cat. No. Name Description Price
HR5-231 Protein Crystallization each $99.00
Completely revised and updated, Protein Crystallization, 2nd Edi-
tion is a greatly expanded follow-up to the best-selling 1st edition.
Completely new chapters on high-throughput methods, mass
spectrometry, microcalorimetry, counterdiffusion, heavy-atom
derivatization, selenomethionine-labeling, rational strategies for
crystallization, and protein modifcation to improve crystallization.
Updated chapters on formulation of the protein before crystalliza-
tion, characterization of the protein by dynamic light scattering,
classic methods and the phase diagram, seeding, and cryoprotection of the crystals. Thirty full-color
plates for evaluating crystallization drops. Separate section of laboratory exercises, ideal for crystalliza-
tion courses. A-Z glossary.
n
Editor: Terese Bergfors
n
Publisher: International University Line;
2nd edition, 2009
n
ISBN-10: 0972077448
n
ISBN-13: 978-0972077446
n
Hardcover: 474 pages
n
Editor: Terese Bergfors
n
Folding 3-D viewer
n
Editor: Terese Bergfors
n
3-D stereo glasses with folding wire legs
for compact storage
d e s c r i p t i o n
PROT EI N CRY STAL L I Z AT I ON
P R A C T I C A L P R O T E I N C R Y S T A L L O G R A P H Y
d e s c r i p t i o n
The Stereopticon is punched from black plastic and folds
completely flat to fit inside a book. The Stereopticon uses plastic
lenses.
S T E R E O P T I C O N S T E R E O V I E W E R
P R A C T I C A L P R O T E I N C R Y S T A L L O G R A P H Y
d e s c r i p t i o n
The Stereo Viewer features an adjustable nose bridge. It is made
of high impact, crystal-clear plastic with a 2.2 magnification, 4.7
focal length and adjustable inter-pupillary distance.
S T E R E O V I E W E R
Order Information
Cat. No. Name Description Price
HR4-517 Stereo Viewer each $12.00
173
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Order Information
Cat. No. Name Description Price
HR4-513 Stereopticon each $9.00
HR4-515 Stereopticon 10 pack $81.00
f e a t u r e s
f e a t u r e s
p r o t e i n c r y s t a l l i z a t i o n s t a n d a r d s
Protein crystal.
Alexey Rak, Max-Planck-Institut fur Molekulare Physiologie, Department of Physical Biochemistry, Dortmund, Germany.
T A B L E O F C O N T E N T S
P A G E S
176 g l u c o s e i s o m e r a s e
177 l y s o z y m e
178 x y l a n a s e
179 l i p a s e b
n
Crystallization grade protein standard
n
Protein standard for crystallization studies and
applications
n
Crystallization training and student demos
Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com 176
a p p l i c a t i o n
f e a t u r e s
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s
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O
HOCH
2
H
HO
OH
H
H
OH
H
OH
O
HOCH
2
OH
H HO
H
OH
HOCH
2
H
+
O
HOCH
2
H
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H
H
OH
H
OH
Glucose Glucose Fructose
Glucose Isomerase catalyzes the isomerization
reaction shown. The reaction is reversible and
goes to equilibrium, approximately equal con-
centrations of fructose and glucose.
d e s c r i p t i o n
Glucose Isomerase can be crystallized by sitting, hanging or sand-
wich drop vapor diffusion, batch, liquid/liquid diffusion, pH shift,
temperature jump, as well as by dialysis. It can be crystallized rapidly
across a broad range of screen conditions which makes the enzyme
an excellent candidate for crystallization studies and demonstrations.
Glucose Isomerase is supplied as a crystal suspension in the follow-
ing medium: Approximately 33 mg/ml in 6 mM TRIS hydrochloride
pH 7.0, 0.91 M Ammonium sulfate, 1 mM Magnesium sulfate.
Glucose Isomerase is prepared by Macrocrystal Oy, exclusively for Hampton Research.
References
1. Carrell, H. L., et al., Proc. Natl. Acad. Sci. USA (1989) Vol 86, 4440-4444.
2. Glucose Isomerase from Streptomyces rubiginosus - potential molecular weight standard for small-angle x-ray scattering. Maciej Kozak. J. Appl. Cryst.
(2005). 38, 555-558.
Order Information
Cat. No. Name Description Price
HR7-100 Glucose Isomerase 1 g $284.00
HR7-102 Glucose Isomerase 100 mg $60.00
G L U C O S E I S O M E R A S E
Glucose Isomerase Crystals
n
Crystallization grade protein standard
n
Protein standard for crystallization studies
and applications
n
Crystallization training and student demos
n
Hen egg white source
a p p l i c a t i o n
f e a t u r e s
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d e s c r i p t i o n
A very cost-effective and convenient crystallization grade lysozyme
(hen egg white). The Lysozyme Kit is a convenient buffer and
lysozyme set for teaching, demonstrations or instrumentation setup
and validation procedures. The kit features 12 vials with 20 mg of
lysozyme and 12 vials of 0.02 M Sodium acetate trihydrate pH 4.6
buffer. To prepare the lysozyme for crystallization, simply add the
supplied buffer to the protein. No weighing, no messing with fluffy
crystalline powder, no mess, and no variability. Each tube can make
up to 1 ml of a 20 mg/ml stock or 200 l of a 100 mg/ml stock. Twelve
sets of tubes and a stable shelf life make this a handy kit to keep in the
fridge. Lysozyme is available separately in an 8 g vial of lysozyme pow-
der. To dissolve the powder, use the Lysozyme Crystallization Buffer.
For teaching and demonstrations requiring crystals in a short period
of time, one should consider the 15-Minute Lysozyme Crystallization
Reagent (30% w/v Polyethylene glycol monomethyl ether 5,000, 1.0 M
Sodium chloride, 0.05 M Sodium acetate trihydrate pH 4.6). For flex-
ibility, one can also use the Lysozyme Crystallization Buffer stock (1.0
M Sodium acetate trihydrate pH 4.6) or the StockOptions Sodium
Acetate buffer kit and 5.0 M Sodium chloride to create custom grid
screens and practicals for demonstrating and teaching the fundamen-
tals of optimization of sample concentration and reagent concentration. The Grid Screen Sodium Chloride
kit is a convenient kit for demonstrating the effect of varying pH and reagent concentration on the solubility
and crystallization of a protein.
Order Information
Cat. No. Name Description Price
HR7-108 Lysozyme Kit 12 x 20 mg plus $60.00
12 x 1 ml solubilization buffer
HR7-110 Lysozyme 8 g $60.00
HR2-731 Lysozyme 1.0 M Sodium acetate trihydrate $29.00
Crystallization Buffer pH 4.6, 100 ml
HR2-805 15 Minute Lysozyme 30% w/v PEG MME 5,000, 1.0 M Sodium $39.00
Crystallization Reagent chloride, 0.05 M Sodium acetate trihydrate
pH 4.6, 100 ml

HR2-637 Sodium Chloride for 5.0 M Sodium chloride, 200 ml $33.00
Lysozyme Crystallization
HR2-233 StockOptions Sodium 1.0 M Sodium acetate trihydrate $195.00
Acetate kit pH 3.6 to 5.6, 21 reagents

HR2-219 Grid Screen Sodium chloride versus pH crystallization $175.00
Sodium Chloride kit screen kit
Lysozyme crystals grown using the Hampton Research Silver Bullets screen
L Y S O Z Y M E
n
Crystallization grade protein standard
n
Protein standard for crystallization studies and
applications
n
Crystallization training and student demos
Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com 178
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Xylanase can be crystallized by sitting, hanging or sandwich drop
vapor diffusion, batch, liquid/liquid diffusion, pH shift, temperature
jump, as well as by dialysis. It can be crystallized rapidly across a
broad range of screen conditions which makes the enzyme an excel-
lent candidate for crystallization studies and demonstrations. It is
supplied as a solution, produced by dissolving pure xylanase crystals
with phosphate buffer and glycerol. The solution is filtered with a
0.2 micron filter and has the final formulation:
Xylanase Concentration: 36 mg/ml
Absorbance at 280 nm: 97
Activity: 648,000 nkat/ml
Glycerol Concentration: 43% w/v
Na/K phosphate: 0.18 M
pH: 7
Specific Gravity: 1.124 g/ml
Xylanase is prepared by Macrocrystal Oy, exclusively for Hampton Research.
References
1. Torronen, A., et al., The EMBO Journal (1994) Vol 13, No 11, 2493-2501.
2. Torronen, A. ,et al., J. Mo. Biol. (1993) Vol 233, 313-316.
3. Crystals of family 11 xylanase II from Trichoderma longibrachiatum that diffract to atomic resolution. Natalia Moiseeva and Marc Allaire. Acta Cryst. (2004).
D60, 1275-1277.
4. Structures of an orthorhombic form of xylanase II from Trichoderma reesei and analysis of thermal displacement. Watanabe et al. Acta Cryst. (2006). D62,
784-792.
5. Cryogenic (<20 K) helium cooling mitigates radiation damage to protein crystals. B. Leif Hanson et al. Acta Cryst (2007) D63, 486-492.
Order Information
Cat. No. Name Description Price
HR7-104 Xylanase 1 g $463.00
HR7-106 Xylanase 100 mg $69.00
X Y L A N A S E
Protein crystals grown using the Silver Bullets screen from
Hampton Research.
Bob Cudney & Peter Nguyen, Hampton Research; Alex McPherson,
University of California, Irvine.
Xylanase Crystals
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Protein crystals grown using the Silver Bullets screen from
Hampton Research.
Bob Cudney & Peter Nguyen, Hampton Research; Alex McPherson,
University of California, Irvine.
Lipase B Crystals
Lipase B Crystals Grown by Macrocrystal Oy
n
Protein standard for crystallization studies
and applications
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Crystallization training and student demos
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d e s c r i p t i o n
Lipase B is a pure, dried, crystalline form of Lipase B from
Candida antarctica produced by submerged fermentation of
a genetically modified Aspergillus oryzae microorganism. It is
a white crystal powder where Lipase B is present as crystals
and there are no other ingredients or buffering salts.
Lipases (EC 3.1.1.3) make up a diverse group of water
soluble enzymes that catalyze the hydrolysis of ester bonds in waterinsoluble, lipid substrates. Lipases
are characterized by their drastically increased esterase activity when adsorbed to a lipid surface (interfacial
activation). Lipases are a form of esterases. They perform essential roles in the digestion, transport, and
processing of dietary lipids such as triglycerides, fats, oils in most, if not all, living organisms.
Chemical name, synonym Triacylglycerol hydrolase EC 3.1.1.3
M
r
35 kDa
pH 5.0 - 7.0
Isoelectric point 6.0
Appearance: White crystal powder
References
1. Crystallographic and molecular-modeling studies of lipase B from Candida antarctica reveal a stereospecifi city pocket for secondary alcohols. Uppenberg, J.,
Ohrner, N., Norin, M., Hult, K., Kleywegt, G.J., Patkar, S., Waagen, V., Anthonsen, T., Jones, T.A. Biochemistry v34 pp. 16838-51, 1995.
2. The sequence, crystal structure determination and refi nement of two crystal forms of lipase B from Candida antarctica. Uppenberg, J., Hansen, M.T., Patkar,
S., Jones, T.A. Structure v2 pp. 293-308, 1994.
3. Crystallization and preliminary X-ray studies of lipase B from Candida antarctica. Uppenberg J., Patkar S.; Bergfors T., Jones, T. A.. Journal of Molecular
Biology, 1994, vol. 235, no2, pp. 790-792.
Order Information
Cat. No. Name Description Price
HR7-099 Lipase B 100 mg $184.00
L I P A S E B
c r y s t a l g r o w t h 1 0 1
Protein crystals.
Kimberly J. Skinner, Structural Biology and Biophysics, Pfizer Global R&D, Sandwich, Kent, United Kingdom.
T A B L E O F C O N T E N T S
P A G E S
182 - 183 p r e l i m i n a r y s a m p l e p r e p a r a t i o n
184 - 185 c r y s t a l g r o w t h t e c h n i q u e s
186 h a n g i n g d r o p v a p o r d i f f u s i o n c r y s t a l l i z a t i o n

187 - 189 s i t t i n g d r o p v a p o r d i f f u s i o n c r y s t a l l i z a t i o n
190 c r y s t a l l i z a t i o n u n d e r o i l
191 - 192 d i a l y s i s c r y s t a l l i z a t i o n
193 v i e w i n g c r y s t a l l i z a t i o n e x p e r i m e n t s
194 b i r e f r i n g e n c e
195 - 196 s a l t o r b i o l o g i c a l c r y s t a l s ?
197
p h v e r s u s n u m b e r o f c r y s t a l s
198
p h p o i n t s t o p o n d e r
198 c r y s t a l g r o w t h v i a p h r e l a x a t i o n
199 p r o t e i n c o n c e n t r a t i o n v e r s u s n u m b e r o f c r y s t a l s

200 - 201 c r y s t a l l i z a t i o n p o l l i n g b o o t h
202 c r y o q u i c k i e s
203 h o w t o : c r y o c r y s t a l l o g r a p h y a n d t h e c r y s t a l c a p s y s t e m
204 u s i n g h a l i d e s f o r p h a s i n g m o l e c u l a r s t r u c t u r e s
205 m i c r o s e e d i n g

206 d e t e r g e n t s a t w o r k
207 - 208 p r o t e i n c r y s t a l l i z a t i o n r e c i p e s
209 - 210 t e m p e r a t u r e a s a c r y s t a l l i z a t i o n v a r i a b l e
212 - 219 t i p s f r o m r a m c
220 - 234 c r y s t a l l i z a t i o n t i p s a - z
235 - 263 c r y s t a l l i z a t i o n t i p s
264 - 265 c r y s t a l l i z a t i o n b u f f e r s t a b l e

266 - 267 s o l u b i l i t y t a b l e
preli mi nary sample preparati on
Lyophilization
Avoid lyophilization. Even though there are many examples of proteins which crys-
tallize after lyophilization (lysozyme, thaumatin, hemoglobin, etc.), lyophilization is
to be avoided when possible. If the protein is lyophilized, it needs to be dialyzed
before crystallization. Dialyze the protein against deionized water or a stabilization
buffer before crystallization. Dialysis will remove non-volatile buffers and other
chemicals that may have been present before lyophilization.
Ammonium Sulfate Precipitation
Avoid using Ammonium sulfate precipitation as a final purification and/or con-
centration step. It is often very difficult to completely remove all the Ammonium
sulfate by a desalting column of dialysis. The remaining trace amounts can inter-
fere with crystallization screening results and create reproducibility problems.
It is not uncommon for trace amounts of Ammonium sulfate in the sample
to cause precipitation or excessive nucleation in screen conditions containing
polyethylene glycol and salt.
Batches
Avoid combining different purification batches for crystallization trials.
Purification conditions and procedures are never identical so each batch should
be screened separately.
Profile the Protein
Ideally, you will purify your own protein, but this is not always reality. So, it is
always a good idea to characterize your protein before beginning crystallization
experiments. Profiling your protein before crystallization can often provide valu-
able clues during screening and optimization of crystallization conditions. Assay
to seriously consider:
SDS-PAGE
Native PAGE or Dynamic Light Scattering
IEF (Isoelectric Focusing) Gel
Mass Spectroscopy
The results of these assays can:
Determine the purity of the sample
Determine the homogeneity of the sample
Identify batch to batch variations
Identify stability problems with the sample
How Pure?
How pure should the protein sample be for crystallization trials? As pure as
possible. That's some answer, is it not? Integrating common sense into the ques-
tion, we might arrive at the following answer. For initial screening, the sample
should be at least 90 to 95% pure on a Coomassie stained SDS-PAGE. Finally, it
does no harm to screen an impure sample as one can always perform further
purification. Remember, crystallization used to be considered a very powerful
purification tool (and still is!).
If the initial screen does not produce crystals, any promising results, or it
becomes next to impossible to improve crystal quality during optimization, one
should consider further purification of the sample.
Storing the Sample
Most proteins can be stored successfully at 4C or -70C.
Check with the person preparing the protein or compare your protein to a simi-
lar protein in the literature for best storage temperature.
Ideally, one should assay the activity and stability of the protein before storage and
then later on at various points in time to determine the sample storage stability.
Repeated freezing and thawing of the sample should be avoided. Aliquot the
sample into multiple small microcentrifuge tubes. Make the aliquots small enough
so that the entire aliquot can be consumed in the experiment after thawing.
Sometimes people like to add Glycerol (10 to 50% v/v) to help proteins bet-
ter tolerate freezing. Avoid this if possible since it is often difficult to remove
Glycerol by dialysis or filtration. The presence of Glycerol is a crystallization vari-
able. It can behave as a precipitant, an additive, or cryoprotectant and therefore
can influence the outcome of a crystallization experiment.
In general, it is better to store proteins more concentrated than diluted. When
too dilute, adsorption of the protein onto the storage container can lead to
significant losses. However, precipitation can sometimes be a problem when the
protein is stored too concentrated.
Sample Handling
Be nice to your protein. Remember that proteins make an excellent food source
for microbes. Protect your sample from microbes by storing the protein at less
than 4C and not leaving it for extended periods of time at room temperature.
When thawing a sample or mixing a lyophilized sample into solution, do not
shake or vortex the protein. Avoid foaming the sample. Foam can be a sign of
denaturation.
Allow the sample to equilibrate to the temperature where the crystallization
experiments will be set up and/or incubated before setting the experiment.
The field of crystal growth is full of opinions and controversy. There are several
opinions on what should be done with the sample just prior to setting the crystal-
lization experiment. Let's have a look at those opinions.
Some like to filter the sample through a 0.2 micron (or smaller, but be sure
to compare the MW of your sample to the pore size of your filter so as not to
stick your sample on the filter) pore size filter into a sterile container. Filtration
can remove microbial contamination (but not the proteases) as well as sample
aggregation. Turbid sample solutions with lots of precipitate should be solu-
bilized or centrifuged before filtration to avoid the ugly experience of sticking
the sample to a filter membrane. Use filters with the smallest possible dead
volume to minimize sample loss. Some of the centrifugal microfiltration devices
are certainly worth consideration. Before filtering the sample, wash or flush the
filter with a small amount of the sample buffer/storage solution. This will test
the filter for compatibility with your sample buffer and remove any trace glycerol
which can sometimes be present from the manufacturer. If possible, test filter
a small aliquot of the sample, and measure the activity/OD before filtering the
entire sample. Do this to test the adherence of the sample to the filter media.
Read and follow the instructions supplied with the filter before introducing the
sample to the filter.
Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com 182 Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com
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P A G E 1 - 2
Some like to centrifuge the sample. Centrifugation removes large sample aggre-
gates and amorphous debris. Post centrifugation views can provide a visual clue
of aggregation/precipitate for seemingly clear solutions. Following centrifugation,
use only the supernatant for crystallization trials.
Others prefer to avoid filtration or centrifugation before setting crystallization
experiments. One view is that the presence of amorphous material or aggregates
can enhance the changes for crystallization by acting as nucleants.
To Azide or Not
Sodium azide (NaN
3
) is an anti-microbial preservative that is sometimes used to pro-
tect samples and crystallization reagents from microbial contamination. Sodium azide
is toxic and should be handled with care. Typical Sodium azide concentrations are 1
mM or if you prefer % measurements, between 0.02% and 0.1% w/v.
If you choose to use Sodium azide remember that:
It is toxic to humans as well as microbes.
It is an inhibitor for some proteins and may become an unintentional ligand
for your sample.
It can interfere with heavy atom derivatization.
Some metal azides are explosive.
There are reports where eliminating Sodium azide from the experiment
improved crystallization.
Alternatives to Sodium Azide include Thymol & Thimerosal.
A final alternative to the use of antimicrobials is the use of proper sterile tech-
nique and materials. Sterile filter all samples and reagents into sterile containers.
Store samples and reagents at 4C or lower. Use sterile pipet tips. Keep your
work area clean. Develop a sterile technique with your crystallization setups.
With common sense, sterile reagents and sample, good technique, and sterile
pipet tips, one can successfully avoid the use of chemical antimicrobials in the
crystallization lab.
Label & Organize Samples
Label samples clearly with the sample identification, batch identification, and date
of storage. Small cryo labels can be very useful here. Color coding samples can
be a nice organization tool. For the sake of easy organization and identification,
it is sometimes more convenient to nest samples. For example, store batches of
small microcentrifuge tubes in 10 ml or 50 ml centrifuge tubes and organize them
by batch or sample.
It is prudent to write down and hold onto detailed notes concerning the puri-
fication, storage, and handling of the sample. It is obvious that one should also
maintain records of crystallization trials which should include:
Sample information
Name of sample
Sample identification (batch, storage location, storage temperature, etc.)
Sample buffer composition, additives, ligands, etc.
Sample concentration
Crystallization experiment information
Method
Drop size and composition
Reagent composition
Temperature
Date
Name of person performing experiment
Questions to Ponder about the Sample
Does a similar sample exist and has it been crystallized?
Does the sample contain free cysteines?
Does the sample contain additives such as Sodium azide, ligands, inhibitors, or
substrates?
Is the protein glycosylated?
Is the protein phosphorylated?
Is the protein N-terminal methylated?
At what temperature is the protein stable?
How does sample solubility and stability change temperature?
How does sample solubility and stability change with pH?
Does the sample bind metals?
Is the protein sensitive to proteolysis?
What class of protein am I working with (antibody, virus, enzyme, membrane
protein)?
What have been the most successful approaches with my class of protein?
What is the source of the sample?
How was the sample purified and stored before it arrived into my hands?
What is in the sample container besides the sample (buffer, additives, etc.)?
Is the sample pooled purification aliquots or a single batch?
How much sample do I have and how much more is available?
How pure is the sample?
How homogeneous is the sample?
Does anyone possess any solubility information on this sample?
What is unique about this protein?
What is necessary chemically and physically to maintain a stable, active sample?
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1991-2009 Hampton Research Corp. All rights reserved. Printed in the United States of America.
This guide or parts thereof may not be reproduced in any form without the written permission of the publishers.
Solutions for Crystal Growth
P A G E 2 - 2
cryst al growt h techni ques
Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com 184 Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com
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There are several techniques for setting up crystallization experiments (often
termed trials), including sitting drop vapor diffusion, hanging drop vapor dif-
fusion, sandwich drop, batch, microbatch under oil, dialysis, and free interface
diffusion. Here we offer an overview of these crystallization techniques.
Sitting & Hanging Drop Crystallization
Sitting and hanging drop methodologies are very popular because they are easy
to perform, require a small amount of sample, and allow a large amount of flex-
ibility during screening and optimization.
Using the sitting drop technique (figure 1), one places a small (0.1 to 40 l)
droplet of the sample mixed with crystallization reagent on a platform in vapor
equilibration with the reagent. The initial reagent concentration in the droplet
is less than that in the reservoir. Over time the reservoir will pull water from the
droplet in a vapor phase such that an equilibrium will exist between the drop and
the reservoir. During this equilibration process, the sample is also concentrated,
increasing the relative supersaturation of the sample in the drop.
The advantages of the sitting drop technique include speed and simplicity. The
disadvantages are that crystals can sometimes adhere to the sitting drop surface
making removal difficult. This disadvantage can turn into an advantage where occa-
sionally the surface of the sitting drop can assist in nucleation. The sitting drop is an
excellent method for screening and optimization. During production, if sticking is a
problem, sitting drops can be performed in the Sandwich Box Setup.
Sitting drop crystallization may be performed using Micro-Bridges

or Glass
Sitting Drop Rods with VDX or Linbro

plates. Both plates can be sealed

with clear sealing tape or plain cover slides for easy viewing and access. Sitting
drop crystallization may also be preformed using the Cryschem Plate. The
Cryschem Plate is a specially designed plate with a post already in the center of
the reservoir. Finally, sitting drop crystallization can also be performed using
one of the 48 or 96 well plates made by Art Robbins Instruments (Intelli-Plate),
Corning (CrystalEX), Douglas Instruments (CrystalClear Strips), Greiner
(CrystalQuick), or Swissci (MRC).
Using the hanging drop technique (figure 2), one places a small (0.1 to 20 l)
droplet of the sample mixed with crystallization reagent on a siliconized glass
cover slide inverted over the reservoir in vapor equilibration with the reagent.
The initial reagent concentration in the droplet is less than that in the reservoir.
Over time the reservoir will pull water from the droplet in a vapor phase such
that an equilibrium will exist between the drop and the reservoir. During this
equilibration process, the sample is also concentrated, increasing the relative
supersaturation of the sample in the drop.
The advantages of the hanging drop technique include the ability to view the
drop through glass without the optical interference from plastic, flexibility,
reduced chance of crystals sticking to the hardware, and easy access to the drop.
The disadvantage is that a little extra time is required for setups.
Sandwich Drop Crystallization
The sandwich drop crystallization method is illustrated in figure 3. The sample
solution that is mixed with the precipitant is placed in the middle of a lower
siliconized glass cover slide. A siliconized glass cover slide is then set in posi-
tion along an upper edge. This allows for a small amount of space between the
cover slides but is close enough so the drop is sandwiched between them. This
technique offers an alternate equilibration method. However, the set up can be
tedious and the plates designed for this method typically have a larger footprint
than most.
Free Interface Diffusion
Free interface diffusion crystallization is used less frequently than sitting or
hanging drop vapor diffusion but it is one of the methods used by NASA in
microgravity crystallization experiments and at least one company has automated
and miniaturized the method. Using this method, one actually places the sample
in liquid contact with the precipitant. When doing so, one attempts to create a
clearly defined interface between the sample and the precipitant. Over time the
sample and precipitant diffuse into one another and crystallization may occur at
the interface, or on the side of high sample/low precipitant or low sample/high
precipitant. The technique allows one to screen a gradient of sample precipitant
concentration combinations. The technique can readily be performed in small
figure 1
Reservoir Solution
figure 3
P A G E 1 - 2
figure 2
H
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O H
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O
[ppt]
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=
[ppt]
reservoir
2
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capillaries (figure 4).
Batch
Batch crystallization is a method where the sample is mixed with the precipitant
and appropriate additives, creating a homogeneous crystallization medium
requiring no equilibration with a reservoir. The technique is popular with small
molecule crystallographers. The advantages to the technique are speed and
simplicity but the disadvantage is that only a narrow space of precipitant/sample
concentration can be sampled in a single experiment. A batch experiment can be
readily performed in a capillary, small container, or plate with a small reservoir.
One must be very close to the conditions which promote crystal growth in order
for this technique to be successful.
Microbatch Under Oil
In this technique, a small drop of the sample combined with the crystallization
reagent is pipetted under a layer of oil. For a true microbatch, the drop is placed
under paraffin oil (figure 5) which allows little to no evaporation, nor concentra-
tion in the drop. A modified microbatch can be performed when the drop is
placed under a mixture of paraffin oil and silicon oil, or straight silicon oil (figure
6). Such oils permit water vapor to permeate from the drop and allow sample
and reagent to concentrate. Unless the drop is equilibrated with a reservoir,
water will leave the drop until only solids remain.
The benefits of crystallization under oil include the use of very small sample and
reagent volumes with less concern for unwanted evaporation, the minimization
of surface interaction with the sample, the ability to precisely control sample and
reagent concentrations during the experiment, and the minimization of conden-
sation during temperature fluctuations.
Dialysis Crystallization
Dialysis crystallization involves placing the sample in a Dialysis Button which is
sealed with a dialysis membrane. Water and some precipitants are then allowed to
exchange while retaining the sample in the cell. The Dialysis Button is placed into
a suitable container holding the precipitant or crystallization media (figure 7). For
example, one might dialyze a sample requiring a high ionic strength for solubility
against a solution of low ionic strength. The technique allows for salting in and
salting out, as well as pH crystallization techniques.
Sample / Reagent Diffusion
Crystal
Sample Reagent
figure 4
Oil
Crystallization
Drop
Oil
Crystallization
Drop
figure 5
Oil
Crystallization
Drop
Oil
Crystallization
Drop
figure 6
Reservoir Solution
Dialysis Button
figure 7
P A G E 2 - 2
Solutions for Crystal Growth
1991-2009 Hampton Research Corp. All rights reserved. Printed in the United States of America.
This guide or parts thereof may not be reproduced in any form without the written permission of the publishers.
hangi ng drop vapor
di ffusi on cryst alli zati on
Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com 186 Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com
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The hanging drop vapor diffusion technique is a very popular method for the
crystallization of macromolecules. The principle of vapor diffusion is straightfor-
ward. A drop composed of a mixture of sample and reagent is placed in vapor
equilibration with a liquid reservoir of reagent. Typically the drop contains a
lower reagent concentration than the reservoir. To achieve equilibrium, water
vapor leaves the drop and eventually ends up in the reservoir. As water leaves
the drop, the sample undergoes an increase in relative supersaturation. Both the
sample and reagent increase in concentration as water leaves the drop for the
reservoir. Equilibration is reached when the reagent concentration in the drop
is approximately the same as that in the reservoir.
Benefits of Hanging Drop Crystallization
Can be cost-effective.
Sample and reagents in contact with a siliconized glass surface.
Easy access to crystals.
Can perform multiple drops (experiments) with a single reservoir.
Good optics for viewing experiments.
Using the VDX Plate
The VDX Plate is a 24 well plate manufactured from clear polystyrene. It is
typically sealed with vacuum grease (HR3-510) and siliconized 22 mm circle
or square glass cover slides. The VDX Plate is also available with sealant which
is a convenient time saver. Rows of the plate are labeled A-D and columns are
labeled 1-6.
1. Apply a bead of vacuum grease along the top edge of the raised reservoir
A1 of the VDX Plate. It is recommended that one apply the vacuum grease
prior to pipetting the reagent. Create a circular bead on the upper edge of
the reservoir. Do not complete the circle. Leave a 2 mm opening between
the start and finish of the circular bead. Apply the cover slide, press to relieve
the air pressure and twist to close the gap. Or simply use the VDX Plate with
sealant. These plates come pregreased.
2. Pipet 1.0 ml of crystallization reagent into reservoir A1 of the VDX Plate.
(Note: Recommended reservoir volume is 0.5 to 1.0 ml)
3. Clean a siliconized 22 mm circle or square cover slide by wiping it with lens
paper and blowing it with clean, dry compressed air. Pipet 1 l of sample
into the center of a siliconized 22 mm circle or square cover slide. (Note:
Recommended total drop volume is 0.1 to 20 l)
4. Pipet 1 l of reagent from reservoir A1 into the drop on the cover slide con-
taining the sample. (Note: Some prefer to mix the drop while others do
not. Proponents of mixing leave the pipet tip in the drop while gently
aspirating and dispensing the drop with the pipet. Mixing ensures a
homogeneous drop and consistency drop to drop. Proponents of not mix-
ing the drop simply pipet the reagent into the sample with no further
mixing.)
5. Holding the cover slide with forceps, the PEN-VAC

, Cover Slide Vacuum

Gadget, or on the edge between your thumb and forefinger, carefully yet
without delay, invert the cover slide so the drop is hanging from the cover
slide.
6. Position the cover slide onto the bead of grease on reservoir A1. Gently press
the slide down onto the grease and twist the slide 45 to ensure a complete
seal.
7. Repeat for reservoirs 2 through 24.
VDX Plate Tips
Note the VDX Plate has a raised cover to protect the cover slides during
transport and storage.
To access a drop and/or reservoir, simply grasp the edge of the cover slide
with forceps or fingertips, twist and pull gently.
VDX Plates can be stacked for convenient storage.
One can pipet multiple drops onto the cover slide. This technique is often
useful when screening additives since one can use the same reservoir with
multiple drops with each drop containing a different additive. This technique
can also be used to screen different drop sizes and ratios versus the same
reservoir. Use care to avoid mixing the drops during pipetting, plate transport,
and plate viewing.
The 0.96 mm Thick glass cover slides or the plastic cover slides are very
durable and most tolerant to rough handling.
Plates for Hanging Drop Crystallization
VDX
VDX with sealant
VDXm
VDXm with sealant
VDX48 with sealant
Modular VDX
Linbro

Greiner ComboPlate
figure 2 Crystallization Droplet
(2 l Sample / 2 l Reagent)
Siliconized cover slide
Solutions for Crystal Growth
1991-2009 Hampton Research Corp. All rights reserved. Printed in the United States of America.
This guide or parts thereof may not be reproduced in any form without the written permission of the publishers.
figure 1
Process of Vapor Diffusion
187
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Solutions for Crystal Growth
si tti ng drop vapor
di ffusi on crystalli zati on
P A G E 1 - 3
The sitting drop vapor diffusion technique is a popular method for the crystal-
lization of macromolecules. The principle of vapor diffusion is straightforward.
A drop composed of a mixture of sample and reagent is placed in vapor equili-
bration with a liquid reservoir of reagent. Typically the drop contains a lower
reagent concentration than the reservoir. To achieve equilibrium, water vapor
leaves the drop and eventually ends up in the reservoir. As water leaves the drop,
the sample undergoes an increase in relative supersaturation. Both the sample
and reagent increase in concentration as water leaves the drop for the reservoir.
Equilibration is reached when the reagent concentration in the drop is approxi-
mately the same as that in the reservoir.
Benefits of Sitting Drop Crystallization
Can be cost-effective.
Can be time-efficient.
Often easier when using detergents, organics and hydrophobic reagents.
Drops can be positioned in a stable sitting position.
Compatible with gels.
Using the Cryschem

Plate
The Cryschem Plate is a 24 well plate manufactured from clear polystyrene. Each
well contains a post in the center which is elevated above the bottom of the res-
ervoir. The smooth, concave depression in the post can hold up to 40 l drops
and the reservoir can hold up to 1.2 ml of reagent. The Cryschem Plate is sealed
with either clear sealing tape or film, or plain 22 mm circle or square glass cover
slides. Rows are labeled A-D and columns are labeled 1-6.
1. Pipet 0.7 ml of crystallization reagent into reservoir A1 of the Cryschem Plate.
(Note: Recommended reservoir volume is 0.5 to 1.0 ml)
2. Pipet 1 l of sample into the post of reservoir A1. (Note: Recommended total
drop volume is 0.1 to 40 l)
3. Pipet 1 l of reagent from reservoir A1 into the drop in post A1. (Note: Some
prefer to mix the drop while others do not. Proponents of mixing leave
the pipet tip in the drop while gently aspirating and dispensing the drop
with the pipet. Mixing ensures a homogenous drop and consistency drop
to drop. Proponents of not mixing the drop simply pipet the reagent into
the sample with no further mixing.)
4. Repeat steps 1 through 3 for the remaining 23 reservoirs.
5. Seal the Cryschem Plate with clear sealing tape or film.
Cryschem Plate Tips
Use Crystal Clear Sealing Tape to seal two rows at a time.
To access a drop and/or reservoir of a Cryschem Plate sealed with tape, simply
make a circular incision in the tape using the inside of the reservoir as a guide.
Use a sharp blade to cut the tape and hold the incised piece of tape with
forceps. The opening can be sealed with another strip of tape or a plain 22
mm circle or square glass cover slide and vacuum grease.
Using Micro-Bridges

The Micro-Bridge is a small bridge (inverted U) manufactured from clear polysty-

rene or clarified polypropylene which contains a smooth, concave depression in
the center of the top region of the bridge (figure 3). The Micro-Bridge can hold
up to 40 l drops. It is inserted into the reservoirs of VDX or Linbro

plates
to perform a sitting drop vapor diffusion experiment. The design is such that the
bridge is quite stable in the reservoir and does
not require the Micro-Bridge to be bonded to
the plate. The Micro-Bridge can be removed
from the plate for crystal manipulation and
observation if desired.
1. Pipet 1.0 ml of crystallization reagent
into reservoir A1 of a VDX plate. (Note:
Recommended reservoir volume is 0.5 to
1.0 ml)
2. Place a clean (blow the Micro-Bridge with clean, dry compressed air before
use) Micro-Bridge into the bottom of reservoir A1 such that the concave
depression in the Micro-Bridge is facing up.
3. Pipet 1 l of sample into the Micro-Bridge in reservoir A1. (Note:
Recommended total drop volume is 0.1 to 40 l)
4. Pipet 1 l of reagent from reservoir A1 into the drop in the Micro-Bridge A1.
(Note: Some prefer to mix the drop while others do not. Proponents of
mixing leave the pipet tip in the drop while gently aspirating and dis-
pensing the drop with the pipet. Mixing ensures a homogenous drop and
consistency drop to drop. Proponents of not mixing the drop simply pipet
the reagent into the sample with no further mixing.)
5. Repeat steps 1 through 3 for the remaining 23 reservoirs.
6. Seal the plate with clear sealing tape or sealant and plain glass cover slides.
Micro-Bridge Tips
To access a drop and/or reservoir sealed with tape, simply make a circular inci-
sion in the tape using the inside of the reservoir as a guide. Use a sharp blade
to cut the tape and hold the incised piece of tape with forceps. The opening
can be sealed with another strip of tape or a plain 22 mm circle or square glass
cover slide and vacuum grease.
Micro-Bridges can be removed for crystal seeding, mounting, manipulation,
and observation.
Micro-Bridges are designed as disposable devices. It is not recommended to
wash and re-use Micro-Bridges.
Micro-Bridges cannot be siliconized or autoclaved.
Crystallization Droplet
(2 l Sample / 2 l Reagent)
figure 2
figure 3
H2O H2O
[ppt]
drop
=
[ppt]
reservoir
2
figure 1
Process of Vapor Diffusion
188
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Using Glass Sitting Drop Rods

The Glass Sitting Drop Rod is a small, solid rod manufactured from clear glass
that has a smooth, concave depression in the center of the top region (figure
4). The opposite end of the glass rod is flat and it can hold up to a 100 l drop.
It is inserted into the reservoirs of VDX plates to perform a sitting drop vapor
diffusion experiment. The Glass Sitting Drop Rod can be secured to the bottom
of the plate using vacuum grease or can be left unattached to the bottom of the
plate. It can be removed from the plate for crystal manipulation and observation
if desired.
1. Pipet 1.0 ml of crystallization reagent into reservoir A1 of a VDX plate. (Note:
Recommended reservoir volume is 0.5 to 1.0 ml; Additional Note: If
vacuum grease will be used to secure
the Glass Sitting Drop Rod to the
plate, apply the grease to the rod
and insert the rod prior to pipetting
reagent into the reservoir.)
2. Place a clean (blow the Glass Sitting
Drop Rod with clean, dry compressed air
before use), siliconized glass rod into the
bottom of reservoir A1 such that the concave depression in the Glass Sitting
Drop Rod is facing up.
3. Pipet 1 l of sample into the Glass Sitting Drop Rod in reservoir A1.
4. Pipet 1 l of reagent from reservoir A1 into the drop in the glass rod A1.
(Note: Some prefer to mix the drop while others do not. Proponents of
mixing leave the pipet tip in the drop while gently aspirating and dis-
pensing the drop with the pipet. Mixing ensures a homogenous drop and
consistency drop to drop. Proponents of not mixing the drop simply pipet
the reagent into the sample with no further mixing.)
5. Repeat steps 1 through 3 for the remaining 23 reservoirs.
6. Seal the plate with two strips of clear sealing tape or a plain glass cover slide
and sealant.
Glass Sitting Drop Rods Tips
To access a drop and/or reservoir of a plate sealed with tape, simply make a
circular incision in the tape using the inside of the reservoir as a guide. Use
a sharp blade to cut the tape and hold the incised piece of tape with forceps.
The opening can be sealed with another strip of tape or a plain 22 mm circle
or square glass cover slide and vacuum grease.
Glass Sitting Drop Rods can be removed for crystal seeding, mounting,
manipulation, and observation.
Siliconize Glass Sitting Drop Rods before use.
Glass Sitting Drop Rods may be washed and used over and over again.
However, if vacuum grease is used to secure the Glass Sitting Drop Rod to the
bottom of the reservoir, we wish you good luck in completely removing the
grease from the rod.
Glass Sitting Drop Rods may be autoclaved.
CrystalClear Strips

CrystalClear Strips consist of a plastic frame and 12 polystyrene strips with a

total experiment capacity of 96 per plate. Each strip contains 8 reservoirs and
platforms. Typical reservoir volumes are 100 l. The smooth, concave depression
on the platform above the reservoir can hold up to a 10 l drop. The CrystalClear
Strip is sealed with clear sealing tape or film.
Pipet 100 l of crystallization reagent in each of the 96 reservoirs. Pipet 1 l of
sample into the depression on the ledge of the first reservoir A1. Pipet 1 l of
reagent from reservoir A1 into the drop on the ledge above reservoir A1. Repeat
steps 2 and 3 for the remaining 95 reservoirs and wells. Seal the CrystalClear
Strips with clear sealing tape or film.
CrystalClear Strips Tips
The strips are available with and without a concave depression for drop place-
ment.
While pipetting reagents into the reservoirs, place a clean pipet tip into the
first empty reservoir. Move the clean tip ahead one empty reservoir with each
reagent addition. This will help one keep track of their pipetting position in
the plate.
Sandwich Box

The Sandwich Box consists of a square polystyrene box, a plastic support, and
a siliconized 9 well glass plate. The Sandwich Box is used when a common
dehydrant system is desired as well as very large drops. Enormous drops can
be pipetted into the siliconized glass wells. The siliconized glass plates offer
excellent optics and can be removed from the plastic box to inspect the drop
for birefringence without optical interference from plastic. Sandwich Boxes
offer unique vapor equilibration kinetics and are very easy to access for crystal
seeding, manipulation, and mounting. The plates are often used for heavy atom
screening and derivatization and are useful for long-term crystal storage when
each well is sealed with a glass slide and vacuum grease.
Open the Sandwich Box and place a plastic support, bottom side facing up into
the box. Apply a bead of vacuum grease to the outer top edge of the box or the
outer lower edge of the lid. Pour 25 ml of crystallization reagent or common
dehydrant into the Sandwich Box. Place the siliconized 9 well glass plate on
top of the inverted plastic support. Pipet the sample into one of the nine wells.
Add the appropriate crystallization reagent to each drop. Place the cover on the
Sandwich Box.
Sandwich Box Tips
Apply a thin bead of vacuum grease around a single depression of the glass
plate and seal the depression with a plain glass cover slide for long term crystal
storage.
Use a siliconized glass depression plate to test a small amount of sample for
solubility with various crystallization reagents.
P A G E 2 - 3
figure 4
Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com
si tt i ng drop vapor
di ffusi on cryst alli zati on
189
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P A G E 3 - 3
Solutions for Crystal Growth
96 Well Plates
The 96 well sitting drop plates offer a variety of drop well configurations and
flexibility in a standard microplate footprint. The 8 x 12 reagent wells in 9 mm
offset can be filled with automated liquid handling systems or manual, single, and
multichannel pipets with a typical reagent fill volume of up to 100 l. The diversity
of the various sample drop wells allow for automated and manual pipetting into
a variety of well shapes, volumes and geometries. Materials range from optically
clear polystyrene to low birefringent polymers. The plates can be manually or
automatically sealed with optically clear sealing tape or film.
Plates for Sitting Drop Crystallization
Cryschem
VDX with Micro-Bridges

or Glass Sitting Drop Rods

Linbro

with Micro-Bridges or Glass Sitting Drop Rods

Greiner ComboPlate with CrystalBridge
Douglas Instruments CrystalClear Strips
Sandwich Box
Intelli-Plate
Corning CrystalEX
Greiner CrystalQuick
Douglas Instruments Vapor Batch
Swissci MRC
1991-2009 Hampton Research Corp. All rights reserved. Printed in the United States of America.
This guide or parts thereof may not be reproduced in any form without the written permission of the publishers.
cryst alli zat i on under oi l
Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com 190 Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com
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1991-2009 Hampton Research Corp. All rights reserved. Printed in the United States of America.
This guide or parts thereof may not be reproduced in any form without the written permission of the publishers.
Method
Crystallization under oil is a method where a small drop of sample combined
with the crystallization reagent of choice is pipetted under a layer of oil. This is
also known as microbatch crystallization.
Oils can also be used as a barrier between the reservoir and the drop in tradi-
tional hanging or sitting drop crystallization experiments. This is known as vapor
diffusion rate control.
Description of Microbatch
The crystallization of proteins
under a thin layer of paraffin
oil was originally described by
Chayen et al. (Appl. Cryst. 23
(1990) 297). In this technique, a
small drop of sample combined
with the reagent of choice is
pipetted under a small layer of
paraffin oil (figure 1). The oil generally used is a mineral oil of branched paraffins
in the C20+ range and allows for little to no diffusion of water through the oil.
Essentially all of the reagents in a batch or microbatch experiment are present
at a specific concentration. Furthermore, there is no significant concentration of
either the protein or the reagent within the crystallization drop.
Description of Modified Microbatch
DArcy et al. (A novel approach to crystallizing proteins under oil, Journal
of Crystal Growth 168 (1996) 175-180) modified the microbatch under oil
technique by using silicone fluids
which are polymeric compounds
composed of repeating dimeth-
ylsiloxane units -(Si(CH
3
)
2
-O-)
n
-.
Using a mixture of 1:1 silicon oil
and paraffin oil, also known as Als
Oil, one can perform a microbatch
experiment under oil and have
diffusion of water from the drop
through the oil, hence a micro-
batch experiment that does allow
for concentration of the sample
and the reagents in the drop (figure 2).
Performing Microbatch / Modified Microbatch
Pipet 6 ml of 100% paraffin oil or 6 ml of 1:1 paraffin/silicon oil (Als Oil) into a
72 Well Microbatch Plate, as shown in figure 1 or 2. Note: One can also utilize
other ratios of paraffin oil and silicon oil to vary the rate of diffusion from
the drop (higher % of silicon oil = more rapid diffusion and evaporation).
Pipet the sample into the appropriate cone-shaped depression in the microbatch
plate followed by addition of the reagent. Typical drop ratios and final drop sizes
are 1:1 and 1 to 2 l. Drops up to 10 l can be achieved under oil using the
microbatch plate. Place plate cover over microbatch plate to prevent dust and
debris from entering experiment. The microbatch method can also be performed
in round bottom, clear 96 well plates. The oil, reagent, and sample can be mixed
quickly and efficiently for fast throughput by using an 8 or 12 channel pipetter.
Description of Vapor Diffusion Rate Control
Chayen (A novel technique to control the rate of vapour diffusion, giving larger
protein crystals J. Appl. Cryst 30 (1997) 198-202) has described a technique
where oils can be used to vary the rate of vapor diffusion. Using mixtures of
paraffin and silicon oil, Chayen reported fewer, larger crystals in the drop.
Using a standard hanging or sitting drop vapor diffusion setup, the drop is first
mixed with reservoir solution, thus preventing oil from entering the drop. Then,
200 l of oil is applied over the reservoir solution (figure 3). The oil acts as a
barrier to vapor diffusion between the reservoir and the drop. Using 100% paraf-
fin oil allows limited amount of vapor diffusion to occur, which in turn causes
the drop to behave like a batch experiment. The drop will then eventually dry
up due to evaporation through the polystyrene plate. Using 100% silicon oil will
give results similar to that when no oil is used. When using a mixture of the
two oils, the rate of vapor diffusion between the drop and the reservoir may
be controlled. The rate of vapor diffusion is also a function of thickness of the
oil layer over the reservoir. Chayen evaluated oil volumes between 100 and
700 l. Oil volumes of 50 to 100 l resulted in crystals similar to the control
without oil. Oil volumes greater than 700 l have a significant delay in the
onset of crystallization, with improved crystal size. Results using hanging drop
were more pronounced than sitting drop which may be due to either surface
effects or the drop geometry in relation to the reservoir which could influence
vapor diffusion kinetics.
Performing Vapor Diffusion Rate Control
Prepare a VDX or Cryschem Plate for a sitting or hanging drop vapor diffusion
experiment. After the reservoir has been added to the drop, pipet between 200
and 700 l of a mixture of paraffin/silicon oil onto the reservoir (figure 3). Seal
the plate.
Plates for Microbatch Crystallization
Douglas Instruments Vapor Batch
Greiner 72 Well Microbatch
Greiner Imp@ct
Swissci MRC Under Oil 96 Well Crystallization Plate
figure 1
figure 2
Paraffin
Oil
Als
Oil
figure 3
Oil Layer
Reservoir
Solution
Solutions for Crystal Growth
191
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Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com
di alysi s crystalli zati on
Solutions for Crystal Growth
Method
Crystallization by dialysis is an easy variation to the typical vapor diffusion
method used to grow crystals. In the dialysis method, the sample in question is
separated from the precipitant by a semi-permeable membrane which allows
small molecules such as ions, additives, buffers, and salts to pass but prevents
biological macromolecules from crossing the membrane. Equilibration kinetics
depend upon the molecular weight cut-off of the dialysis membrane, the precipi-
tant, the ratio of the volume, the concentration of the components inside and
outside of the dialysis cell, and the geometry of the cell.
Description
The Dialysis Buttons offered by Hampton Research are machined from
transparent perspex. The button has a chamber which varies from 5 to 350 l
depending upon which size button
one chooses to use. The sample
is placed in this chamber so as to
create a slight dome of liquid at the
top edge of the button. A dialysis
membrane (having the appropriate
molecular weight cut-off) is placed
over the top of the button/sample
and is held in place with an O-ring.
The O-ring is held in place by a groove in the button. Dialysis Buttons are notori-
ously tricky to set up since beginners often trap air bubbles between the sample
solution and the membrane which impedes dialysis. With a little practice using
a golf tee or applicator, one can master the technique. Dialysis Buttons are
supplied with O-rings and a golf tee. Dialysis membrane discs and applicators
are available separately. The applicator is used to apply the membrane and the
O-ring to the buttons.
Using the Dialysis Button
A typical dialysis experiment is used to take the sample from the presence of
a high ionic strength solution to a lower ionic strength solution. However, the
technique can just as easily be used to proceed from low ionic strength to a
higher ionic strength. This is accomplished by placing the sample in high ionic
strength into the Dialysis Button, sealing the button with a dialysis membrane,
and placing the sealed button in a solution of ionic strength lower than that
inside the button. Salts, ligands, and compounds smaller than the pore size
of the dialysis membrane will leave the button as long as their concentration
is lower on the opposite side of the membrane. Once the concentration of the
diffusible species is the same on both sides of the membrane, the system is in
equilibrium.
Cleaning
The buttons can be cleaned with soap and deionized water. Do not clean them
with organic solvents as this may turn the perspex opaque.
Practical Example
The following two practicals offer examples of how to set up a dialysis experi-
ment.
Practical 1 - Carboxypeptidase A
1. Using Carboxypeptidase A (Sigma-Aldrich CO386
or CO261), make an 8 to 20 mg/ml solution of the
Carboxypeptidase A in 20 mM Tris HCl pH 7.5, 1.5
M LiCl.
2. Place 100 ul of 10 mg/ml carboxypeptidase in 20
mM Tris HCl, 1.5 M LiCl, pH 7.5 in a 100 ul Dialysis
Button. The droplet should have a slight dome
shape following the hemispheric edge of the top
of the button.
3. Seal the button with dialysis membrane. Using a
one inch (2.5 cm) square of dialysis membrane
which has equilibrated in water, place the mem-
brane over the top of the button. Place an inverted
golf tee on top of the membrane and button. Roll
the O-ring down the applicator until the O-ring
rolls off onto the edge of the button. Roll the O-ring into the machined groove
on the edge of the button. Remove the applicator. There should be no bub-
bles between the membrane and the sample inside the button. Bubbles will
prevent dialysis. Some researchers prefer to use the rubber tipped plunger of
a syringe with a modified syringe body to apply the O-ring. Others prefer to
use an applicator. Try each and see which method works best.
4. Place 0.9 ml of 20 mM Tris HCl, pH 7.5 in the reservoir of a VDX Plate, a
Linbro

Plate, or small chamber which can be sealed.

5. Place the Dialysis Button in the well, membrane side up. Be sure the reservoir
solution covers the top of the membrane/button. Seal the VDX plate using
grease and a cover slide.
6. Observe under a microscope. Crystals will appear within 2 to 3 days. Final
concentration of LiCl will be 0.15 M.
Reference for the above protocol: Dr. Jim Pflugrath and Dr. Gary Gilliland, Cold Spring Harbor Laboratory
Protein Crystallography Workshop.
O-Ring
Golf Tee
Dialysis Button
Sample Chamber
O-Ring Support
figure 1
figure 2a
Cover Slide (or Sealing Tape)
Well of VDX
Crystallization Plate
Reservoir Solution
Dialysis Button
Vacuum
Grease
figure 3
P A G E 1 - 2
figure 2b
Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com 192 Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com
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1991-2009 Hampton Research Corp. All rights reserved. Printed in the United States of America.
This guide or parts thereof may not be reproduced in any form without the written permission of the publishers.
Practical 2 - Lysozyme
1. Prepare 10 mg/ml lysozyme in 50 mM Sodium acetate trihydrate pH 4.5. Filter
the solution using a 0.2 micron filter.
2. Fill a 100 l Dialysis Button with 100 l of the lysozyme solution as described
for the carboxypeptidase practical.
3. Pipet 1 ml of 50 mM Sodium acetate trihydrate buffer into a small (5 ml) bea-
ker.
4. Place the filled button, membrane side up in the beaker.
5. Pipet a small amount of concentrated Sodium chloride into the beaker such
that the final concentration of Sodium chloride in the beaker is 0.2 M. Seal the
beaker with parafilm and store at room temperature.
6. Increase the concentration of Sodium chloride each day by 0.2 M. Repeat until
crystals are observed in the button.
Reference for the above protocol: Crystallization of nucleic acids and proteins, a practical approach. Edited by A.
Ducruix and R. Giege, Oxford University Press, 1992.
Considerations
Just as in a vapor diffusion experiment, the path is often as important as the
endpoint in a dialysis experiment. The path is the equilibration course which the
solution inside and outside the button take toward achieving equilibrium. This
course can be changed by manipulating the following:
Ratio of button volume/reservoir volume
Button and reservoir components & concentration
Molecular weight cutoff of dialysis membrane
Viscosity of solutions
Plus the usual assortment of crystallization variables
including pH, sample concentration, temperature, etc.
Variations of Dialysis
Macrodialysis
The sample is loaded into dialysis tubing of the appropriate molecular weight
cutoff and is dialyzed against the appropriate reservoir solution. This method
typically requires at least 100 l of sample and can be performed with liters of
sample in large dialysis tubing.
Zeppenzauer Cells
Capillary tubes are closed with dialysis tubing or gel plugs. See Zeppenzauer, M.
1971, Methods In Enzymology, 22, 253.
Microcap Dialysis
The sample is placed in a glass capillary with one end sealed with wax, the other
with dialysis membrane. The tube is placed in a microcap/small centrifuge tube
filled with the appropriate reagent. See Crystallization of nucleic acids and pro-
teins, a practical approach, Edited by A. Ducruix and R. Giege, Oxford University
Press, 1992.
Double Dialysis
This method reduces the rate of equilibration and can provide enhanced control
over the crystallization of the sample. Simply put, a Dialysis Button is prepared
and placed inside a reservoir sealed with a dialysis membrane, which is in turn
placed inside another reservoir. Confused? See Thomas, D.H., et al., 1989,
Protein Engineering, 2, 489.
References and Readings
1. Crystallization of nucleic acids and proteins, Edited by A. Ducruix and R. Giege, The Practical Approach Series, Oxford
Univ. Press, 1992.
2. Preparation and analysis of protein crystals. McPherson, A. Eur. J. Biochem. 189, 1-23, 1990.
3. Zeppenzauer, M. et al., Crystal. of horse liver alcohol dehydrogenase complexes from alcohol solutions. Acta Chem Scan,
21, 1099, 1967.
4. Christopher Bunick, A.C.T. North, and Gerald Stubbs., Evaporative microdialysis: an effective improvement in an estab-
lished method of protein crystallization. Acta Cryst. D56, 1430-1431, 2000.
Solutions for Crystal Growth
P A G E 2 - 2
di alysi s cryst alli zati on
Single Crystal
Clear Drop
Skin/Precipitate
Precipitate
Precipitate/Phase
Quasi Crystals
Microcrystals
Needle Cluster
Plates
Rod Cluster
figure 1
Typical observations in a crystallization experiment.
193
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Solutions for Crystal Growth
Observing the Experiment
Gently set the plate onto the observation platform. If the
platform is smooth and free of protrusions, one may simply
slide the plate in the X and Y directions on top of the viewing
platform to view each of the drops. Use low magnification to
view and center the drop in the field of view. Scan the drops
at 20 to 40x magnification. When something suspicious
appears, increase the magnification to 80 or 100x for a better
view. Scan the entire depth of the drop using the fine focus
control of the microscope. Sometimes crystals will form at
different depths of the drop because different areas of the
drop can equilibrate at different rates. Also, crystals some-
times form at the top of a drop and as the crystal gains mass,
it will fall to a lower portion of the drop. Scrutinize every-
thing until you are familiar with the differences between
crystals, microcrystals, precipitate, and sweater fuzz. True
crystals will feature edges. Precipitate does not have edges.
Crystals can appear as needles, blades, walnuts, spherulites,
plates, and various geometric shapes. Crystals vary in size
anywhere from a barely observable 20 microns to 1 or more
mm but most seem to fall in the range of 0.05 to 0.5 mm.
Figure 1 shows typical examples of what one might observe
in a crystallization experiment.
Diffractable Crystal
Crystals useful for x-ray diffraction analysis are typically
single, 0.05 mm or larger, and free of cracks and defects.
Differentiating Between Microcrystals & Precipitate
Microcrystals (less than 0.02 mm) can be difficult to dif-
ferentiate from precipitate, especially under low power or
with a low to medium quality microscope. Differentiate
microcrystals from amorphous precipitate by looking for
birefringence (light colored shiny spots under a polarizer in
dark field mode = crystals). Other tests to differentiate crys-
tals from precipitate include streak seeding, or the use of a
small amount (1 l) of colored, low molecular weight, water
soluble dye (crystal violet and methylene blue will often pen-
etrate the solvent channels of macromolecular crystals and
color them, where as precipitate will not be colored).
Precipitate can appear as clumps, fine wispy clouds, or
anything in between and can range in color from white
to yellow, brown, or rust. During screening and very pre-
liminary optimization, one may wish to observe the drops
immediately after setup, one day later, and each day there-
after for the first week. Observations may be performed
once a week thereafter until the drops turn into a crust of
deceased sample and reagent. Never throw plates away until
the drop is dead. Why? Most crystallization plates are made
from polystyrene which allows for some evaporation over
time. Evaporation leads to increased relative supersaturation
and over time, may also lead to an increase in crystals. Time
also can lead to changes in the protein (denaturation of less
stable forms, proteolytic cleavage, and other changes) which
might promote crystallization. Take careful notes during
observations and be especially conscious of changes that
occur between observations. Most crystallization observa-
tions are done at room temperature since this is where one
will find most microscopes and it is most comfortable.
Cold Experiments
4C experiments may be observed by moving the micro-
scope into a cold room. Allow time for the microscope
to equilibrate to 4C to prevent fogging of the optics as
well as unnecessary temperature transfer from the warm
microscope to the cold experiment. Wear a warm jacket
with gloves to stay as comfortable as possible in the cold
room. Excessive moisture in a cold room can be very
destructive to a microscope so check with your mainte-
nance group to keep the cold room as dry as possible. If a
cold room is unavailable, one is forced simply to work fast,
moving plates from an incubator to the microscope care-
fully and making rapid, yet thorough observations. Move
only one plate at a time and gently close the incubator
door between transfers since slamming the door will cause
vibrations which can influence crystallization. Cold experi-
ments tend to fog up rapidly, especially if the light source
is hot (if no infrared filters or light pipes are used). This
is difficult to avoid and is one reason researchers prefer
working in cold rooms.
1991-2009 Hampton Research Corp. All rights reserved. Printed in the United States of America.
This guide or parts thereof may not be reproduced in any form without the written permission of the publishers.
The technical mumbo jumbo first. The physical properties of isotropic materials,
such as glasses, liquids, and amorphous materials do not depend on direction.
However, most properties of a wide variety of crystals (including liquid crystals)
do show such variation. This anisotropy of physical properties originates in the
anisotropic build-up of the materials (crystal structure). Anisotropy in the optical
properties of uniaxial crystals is referred to as either birefringence or dichroism,
depending on whether the index of refraction or the absorption coefficient is con-
cerned. Birefringence means that there are two distinct speeds with which light
can propagate, depending on the direction of propagation. When a light ray splits
into two beams as it passes through a material, the effect is called birefringence
(or double refraction) and the material is birefringent. If you look at something
through a birefringent material, youll see double. The word birefringence comes
from the Latin bi- (twice) plus refringere (to break up). Thus, the light rays are
broken in two by a birefringent material. One well-known example of a birefrin-
gent medium is crystalline calcite (calcium carbonate). If you look at the world
through a clear crystal of calcite (calcium carbonate), you will see double. Place
such a crystal on a drawing, and you'll see two overlapping copies of the drawing.
The molecular structure of calcite causes double refraction, in which each light ray
is split into two rays that emerge from the crystal at slightly different angles. Calcite
shows this more clearly than most crystals, but quartz and many other crystalline
minerals also split light ray.
Now, the practical interpretation for crystal growers. You might hear the word
birefringence used quite often by crystal growers when viewing crystals under a
microscope. Here, crystal growers are stretching the definition of the term bire-
fringence to describe the colorful display produced by biological macromolecular
crystals when polarized light is passed through the crystal. See figure 1.
A light microscope with polarizing optics is required to observe birefringence. The
following path is a typical setup where light passes from the light source through
the first polarizing lens, next the specimen (crystal), then the second polarizing
optic, finally the magnifying optics and into your eye. On many typical polarization
setups, the second polarizing filter can be rotated while the specimen is stationary.
Rotating the polarizing optic without something to rotate the plane of polarized
light in the path (i.e. a crystal) will result in one seeing light, dark, light, dark, as
the filter is rotated. But if a crystal with birefringent properties (i.e. a biological
macromolecular crystal) is positioned in between the two polarizing filters, one
will observe changing colors as the polarizing filter is rotated. Specifically, when
the polarizing filters are aligned such that the field is dark, a birefringent object
(crystal) will glow with color.
Birefringence is one way we can differentiate amorphous precipitate from
microcrystals in a drop when viewed under a microscope. Precipitate does not
have birefringent properties while most biological macromolecular crystals do.
One drawback with using birefringence in today's crystal growth world is that most of
the crystallization devices utilized are made from plastic such as polystyrene and poly-
propylene. These plastics are optically active and can be birefringent. In fact, often times
the colors we see displayed in crystals are contributions from the birefringent plastic.
See figure 2. However, it is still possible to observe microcrystalline birefringence in the
plastic trays, but there is usually a contributory effect from the plates used to grow the
crystals. One way to avoid this is to grow crystals in a glass device or at least observe the
crystals in a path that is free of birefringent plastics. Some crystallization plates are avail-
able in a low birefringent material.
Birefringent precipitates will glow, sparkle, or glisten. See figure 3.
To test for birefringence, position the polarizers so the field of view is dark
WITHOUT the crystallization plate or setup. Place the tray into position on the
microscope. If a crystal is birefringent, some of the light passing through the crys-
tal will be rotated and pass through the second (analyzing) polarizing filter. The
intensity of the transmitted light will increase and decrease as the crystal or the
polarizer is rotated. Remember, birefringence is NOT ALWAYS clearly visible when
polystyrene or polypropylene is in the light path (i.e. when you use plastic slides or
crystallization plates). However, a birefringent crystal viewed in most plastic trays
or plastic cover slides will have a different color than the background (i.e. plastic
plate) or foreground (plastic slide). Finally, birefringence is a property of crystals,
both biological (proteins, peptides, and nucleic acids) and inorganic crystals (salts).
Birefringence is MORE pronounced in inorganic crystals (salt).
A quick comment on what to do with birefringent precipitates. Streak seeding is
a common and often successful method of taking advantage of microcrystalline
precipitate to grow large single crystals.
figure 2
figure 1
figure 3
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1991-2009 Hampton Research Corp. All rights reserved. Printed in the United States of America.
This guide or parts thereof may not be reproduced in any form without the written permission of the publishers.
Solutions for Crystal Growth
bi refri ngence
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Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com
Solutions for Crystal Growth
P A G E 1 - 2
First of all, before deciding on how to distinguish whether the crystal is inorganic
(salt) or biological (protein), take a picture of the crystal before you destroy it
or make it disappear.
The most definitive test is to obtain an x-ray diffraction pattern of the crystal. A
diffraction pattern of a protein crystal may look like that in figure 1. Want to take
a less direct approach? Read on.
The following are ways to differentiate a biological crystal from an inorganic crystal.
1. Dehydration
A biological crystal typically has very significant solvent content and will dehydrate
when removed from the drop or when the drop is allowed to evaporate from
around the crystal (figure 2). Inorganic crystals typically do not possess large
solvent channels and have very little solvent content. Removing an inorganic
crystal from the drop or allowing the drop to evaporate from around the crystal
will typically not destroy or change the appearance of the inorganic crystal.
2. Physical Manipulation
A biological crystal behaves more like an ordered gel than a hard crystal and will
powder, crumble, or break easily when touched with a probe such as a Micro-
Tool or needle (figure 3). Inorganic crystals can also break apart, but they require
more force and typically make a click or crunching sound when breaking apart
under the force of a probe. The inorganic crystals are typically more dense than
protein crystals and once broken, the pieces fall quickly and stay put on the
bottom of the drop.
3. Birefringence
A biological crystal, unless it is cubic, will be weakly birefringent under cross
polarizers (figure 4). Inorganic crystals are typically strongly birefringent under
cross polarizers. Some plastic plates and materials are also birefringent so this
test is more easily performed and interpreted in an all-glass environment or in a
plate made from a low birefringent plastic.
4. Dye absorption
A biological crystal typically has large solvent channels which will accommodate
a small molecule dye. Small molecule dyes can travel into these solvent channels
and color the crystal (figure 5). Inorganic crystals do not possess such solvent
channels and will not absorb the small molecule dye. Dyes are chemicals and
have solubility limits. So it is possible that a crystal grown in a reagent of high
relative supersaturation may have a reagent concentration that will precipitate
or even crystallize the dye. Most dyes under such conditions will crystallize into
needles or whiskers and of course be colored. So before adding dye, take a
picture or memorize the location of crystals in the drop in case the crystal itself
forms crystals. Finally, diluting the drop with dye can sometimes decrease the
relative supersaturation in the drop to the point where the biological crystal will
dissolve. Once the drop equilibrates again with the reservoir, the crystal may
reappear and it may appear in a location different from the original crystal. For
dyes, consider Izit (HR4-710) from Hampton Research.
5. Control Experiment
Using the same sample buffer, same reagent, same volumes and same hardware,
set an experiment identical to the one that produced the crystals, except leave
the sample out of the experiment. Simply replace the sample with the sample
buffer. Dont get lazy and use water instead of sample buffer because the sample
buffer may be a variable in the formation of the crystals. If a crystal forms that
appears visually similar to the original crystal you likely have an inorganic crystal
in the original setup.
6. Run a Gel
Collect, wash, and dissolve the crystals. Run the sample on SDS-PAGE. If a band
indicates the presence of your sample, there is a high probability that your origi-
nal crystals are biological.
Hampton Research would like to thank Joe Luft at the Hauptman Woodward
Medical Research Institute for his help in putting together the information about
inorganic and biological crystals.
figure 1
X-ray diffraction pattern from a protein crystal.
figure 2
Below is a happily hydrated protein crystal. On the next page is an unhappy,
dehydrated protein crystal, removed from the drop.
salt or bi ologi cal crystals?
Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com 196 Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com
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1991-2009 Hampton Research Corp. All rights reserved. Printed in the United States of America.
This guide or parts thereof may not be reproduced in any form without the written permission of the publishers.
Dehydrated
Protein Crystal
Before: Pre-Crush
After: Crush
figure 3
A protein crystal before and after being bullied by a probe.
figure 5
Protein crystal stained with a dye.
figure 4
A weakly birefringent biological crystal.
Solutions for Crystal Growth
P A G E 2 - 2
salt or bi ologi cal crystals?
197
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Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com
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250
200
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0
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2
2.5
3
3.5
4
4.1
4.2
4.3
4.4
4.5
4.6
4.7
4.8
4.9
5
5.1
5.2
5.3
5.4
5.5
5.6
5.7
5.8
5.9
6
6.1
6.2
6.3
6.4
6.5
6.6
6.7
6.8
6.9
7
7.1
7.2
7.3
7.4
7
.5
7
.6
7
.7
7
.8
7
.9
8
8
.1
8
.2
8
.3
8
.4
8
.5
8
.6
8
.7
8
.8
8
.9
9
9
.1
9
.2
9
.3
9
.4
9
.5
9
.6
9
.7
9
.8
9
.9
1
0
1
0
.1
1
0
.2
1
0
.3
1
0
.4
1
0
.5
1
0
.6
1
0
.7
1
0
.8
1
0
.9
1
1
1.0 1
1.5 1
2.0 0
2.5 1
3.0 7
3.5 17
4.0 46
4.1 5
4.2 32
4.3 11
4.4 12
4.5 78
4.6 39
4.7 10
4.8 21
4.9 16
5.0 102
5.1 8
5.2 51
5.3 28
5.4 37
5.5 98
5.6 53
5.7 21
5.8 23
5.9 24
6.0 239
6.1 9
6.2 54
6.3 24
6.4 29
6.5 217
pH Number of Crystals
6.6 38
6.7 46
6.8 116
6.9 35
7.0 379
7.1 21
7.2 78
7.3 32
7.4 77
7.5 251
7.6 45
7.7 13
7.8 49
7.9 6
8.0 220
8.1 3
8.2 15
8.3 14
8.4 18
8.5 79
8.6 7
8.7 8
8.8 8
8.9 3
9.0 32
9.1 2
9.2 2
9.3 2
9.4 1
9.5 19
9.6 0
9.7 1
9.8 8
9.9 0
10.0 7
10.1 0
10.2 1
10.3 0
10.4 0
10.5 2
10.6 0
10.7 0
10.8 0
10.9 0
11.0 1
pH Number of Crystals
pH Number of Crystals
pH versus number of crystals grown for 2,953 biological
macromolecules reported from the BMCD.
Solutions for Crystal Growth
1991-2009 Hampton Research Corp. All rights reserved. Printed in the United States of America.
This guide or parts thereof may not be reproduced in any form without the written permission of the publishers.
crystal growth vi a
p
h relaxati on
Try the following if sample solubility is pH dependent. Determine a pH and salt
concentration where the protein is insoluble. A simple way to do this, if sample
supply is limited, is to review old drops and look for those which contain pre-
cipitate. Review the data and select the reagent, reagent concentration, and pH
most often associated with precipitate. Or, if sample is abundant, using a micro-
scope, a microscope slide, or cover slide, empirically perform solubility tests by
adding 1 l of protein to 1 l of buffer (and/or precipitant) and watch the drop
for the formation of precipitate within a minute or so (not too long or the drop
will evaporate and you will always observe precipitate!). Once reagent and pH
points have been determined to promote a sample solubility minima, set hang-
ing or sitting drop vapor diffusion experiments with a smattering of the solubility
minima conditions. Once the sample has precipitated, add acetic acid (acidic) or
ammonium hydroxide (basic) to the drop (try 1 l of 0.1 M stock and adjust the
amount and/or concentration of the stock accordingly) until the precipitate is
redissolved. Seal the reservoir. The addition of the acid or the base will, in most
experiments, solubilize the precipitate. Since acetic acid and ammonium hydrox-
ide are volatile, the acidic/basic reagent will leave the drop. As the acidic/basic
reagent leaves the drop, the pH of the drop will relax to approximately the original
pH prior to the addition of the acidic/basic reagent. As the relaxation approaches
the original pH, the protein will approach a point of solubility minima and here is
when there is the chance for crystallization to occur. Variables to consider when
refining the pH relaxation technique include initial pH, buffer concentration, buf-
fer type, precipitant type, precipitant concentration, concentration and volume
of acetic acid/ammonium hydroxide added to the drop, reservoir volume, and
drop volume. One can also incorporate additives (organic solvents, chaotropes,
detergents, ions, salts, etc.) into the procedure to evaluate the effect of the addi-
tive on sample-sample and sample-solvent interactions. Sound tedious? Well, its
not as bad as it sounds. Approach the technique as both a crystallization experi-
ment and a crude solubility measurement. If no crystals are grown using the
method, all is not lost. One has at least obtained a great deal of pH and reagent
dependent solubility data which can be used to design additional crystallization
experiments. What should one do with the old experiments full of precipitate
and no crystals? Move the plates to warmer and then cooler temperatures. Note
changes in sample solubility. See a difference? Consider temperature relaxation
as another crystallization method.
p
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Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com 198 Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com
Solutions for Crystal Growth
n
Never use a pH probe containing AgCl when measuring the pH and/or titrat-
ing the pH of Tris and Tris HCl buffers since the AgCl will complex with the
Tris which can lead to erroneous pH readings. Use a Tris-compatible pH
probe when adjusting the pH of Tris buffers.
n
Follow the manufacturers recommended use and maintenance instructions
to keep the pH probe accurate and precise.
n
pH measurements are only as good as the probe.
n
Glass electrode probes are more precise and accurate than gel-filled plastic
probes.
n
Calibrate pH probes each and every day. Refresh calibration standard solutions
frequently to avoid contamination and evaporation.
n
A two-point calibration with the pH to be measured sandwiched between
the calibration points is better than a single or triple point calibration.
n
Bacterial contamination of pH standards, storage solution, and water can
cause pH fluctuations in reagents.
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1991-2009 Hampton Research Corp. All rights reserved. Printed in the United States of America.
This guide or parts thereof may not be reproduced in any form without the written permission of the publishers.
1991-2009 Hampton Research Corp. All rights reserved. Printed in the United States of America.
This guide or parts thereof may not be reproduced in any form without the written permission of the publishers.
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Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com
Solutions for Crystal Growth
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7
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1
Num
ber of Crystals
500
450
400
350
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250
200
150
100
50
0
0
50
100
150
200
250
300
350
400
450
500
1 - 20 mg/ml
40
39
38
37
36
35
34
33
32
31
30
29
28
27
26
25
24
23
22
21
120
100
80
60
40
20
0
0
20
40
60
80
100
120
N
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rystals
21 - 40 mg/ml
60
59
58
57
56
55
54
53
52
51
50
49
48
47
46
45
44
43
42
41
30
25
20
15
10
5
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0
5
10
15
20
25
30
N
um
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rystals
41 - 60 mg/ml
80
79
78
77
76
75
74
73
72
71
70
69
68
67
66
65
64
63
62
61
7
6
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4
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1
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0
1
2
3
4
5
6
7
N
um
ber of C
rystals
61 - 80 mg/ml
1 23
2 76
3 80
4 97
5 392
6 78
7 60
8 86
9 27
10 483
11 18
12 35
13 21
14 14
15 127
16 15
17 10
18 16
19 3
20 159
21 3
22 5
23 4
24 1
25 51
26 0
27 5
28 3
29 3
30 110
31 0
32 2
[Protein] Number of Crystals
33 3
34 5
35 12
36 2
37 1
38 0
39 1
40 42
41 0
42 2
43 0
44 0
45 0
46 1
47 0
48 1
49 0
50 27
51 0
52 2
53 0
54 0
55 1
56 0
57 0
58 0
59 0
60 30
61 0
62 0
63 0
64 0
65 0
66 0
67 0
68 0
69 0
70 7
71 0
72 0
73 1
74 0
75 1
76 0
77 0
78 0
79 0
80 4
Number of Crystals
Number of Crystals
Protein concentration (mg/ml) versus number of crystals grown for 2,953
biological macromolecules reported from the BMCD.
[Protein]
[Protein]
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cryst alli zat i on polli ng booth
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Answer Percentage
4C only 3%
25C only 43%
4C and 25C only 33%
4C and 25C and other temperatures 21%
What temperatures do you routinely use for crystallization?
Answer Percentage
Very important
4%
Important 35%
45%
Neither important nor unimportant 7%
Slightly important
Not important at all
8%
How important is it to keep your chosen crystallization temperature constant
during the entire crystallization experiment (e.g. during crystal growth and
crystal inspection)?
Answer Percentage
Hanging Drop Vapor Difusion
Sitting Drop Vapor Difusion
Microbatch
Free Interface Difusion
Dialysis
31%
0%
0%
3%
What crystallization method do you use most often for crystal optimization?
66%
Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com
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Answer Percentage
Hanging Drop Vapor Difusion
Sitting Drop Vapor Difusion
Microbatch
Free Interface Difusion
Dialysis
36%
0.5%
0.5%
What crystallization method do you use most often for screening?
58%
5%
Answer Percentage
Cleave it (remove His tag) and screen
Leave tag intact and screen
Leave it & screen. No xtals, cleave & screen.
Based on your own experience, if your protein has a His tag, do you cleave
it or leave it when you screen for crystallization conditions?
59%
15%
26%
1991-2009 Hampton Research Corp. All rights reserved. Printed in the United States of America.
This guide or parts thereof may not be reproduced in any form without the written permission of the publishers.
Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com 202 Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com
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Solutions for Crystal Growth
cryo qui cki es
Cryobuffer
Weve had good success using the well solution directly as the foundation of a
cryobuffer in several situations where crystals cannot be grown directly in the pres-
ence of cryoprotectant, and where crystals don't tolerate transfer to artificial mother
liquors. The basic protocol is as follows:
1. Remove 100 l of the well solution after crystals have grown.
2. Split this sample into two 50 l aliquots.
3. Add 7.5 mg of Dextrose (glucose) to the first aliquot and 15 mg of Dextrose to
the second. Dissolve by gently pipetting with a wide-bore tip. This will give
two sequential well solutions that now contain 15% and 30% w/v Dextrose. If
all the dextrose won't go into the second aliquot, spin hard and remove the
supernatant.
4. Transfer the crystal to aliquot 1, equilibrate for 3 minutes, then to aliquot
number 2, then freeze.
We've had a few crystals that routinely crack or blow up when transferred to
artificial mother liquor that behave well when transferred to well solution plus
glucose. We assume that there is some aspect of the crystal drop (pH, ionic ten-
sion, precipitant concentration) that is more effectively reproduced within the
well than by separately prepared mother liquors.
The nice thing about the protocol above is that you don't get much of a volume
increase when dry dextrose is dissolved in the well solution, so the components
in the solution are not diluted.
Finally, if you don't get a really good freeze, you can try to add about 5% v/v
Glycerol to aliquot 2 in addition to the 30% w/v Dextrose.
Thank you Barry Stoddard for sharing this tip.
Cryo Tip
When adding glycerol to crystallization experiments for cryogenic data collec-
tion, remember that some proteins are more soluble in glycerol and one might
have to increase the concentration of the primary precipitant (Example: Increase
Ammonium sulfate from 2.0 M to 2.1 M or 2.2 M in the presence of 20% v/v
Glycerol).
Cryo-Soaking Method
Eom et al. have reported improved diffraction using a novel cryo-soaking
method. Crystals were grown using the following condition: 1.8 M Ammonium
sulfate, 0.1 M Sodium citrate pH 5.1 (hanging drop, 0.5 ml reservoir). To improve
diffraction quality, the crystals were sequentially transferred to cryosolvents con-
taining 5 to 25% v/v Glycerol in five steps (5, 10, 15, 20, and 25%). Equilibration at
each step was 30 seconds. Note that the cryoprotectant (glycerol) also contains
the mother liquor used to grow the crystals. Equilibration at each step was 30
seconds. After the final transfer, the crystals were equilibrated for 30 minutes.
The cryosolvent containing the crystal(s) was then air dried until Ammonium
sulfate crystals began to appear. The authors report this took about 20 minutes at
room temperature. Diffraction resolution using the novel cryo-soaking method
improved from 3 angstrom to 2.2 angstrom. (Editors Note: Yet another case of
crystal dehydration?)
Reference:
Acta Cryst (1999) D55 Pages 1601-1603 Crystallization and preliminary X-ray crystallographic studies of the D2 region of the
skeletal ryanodine receptor. Seha Kim, Dong Wook Shin, Do Han Kim and Soo Hyun Eom. E-mail: eom@kjist.ac.ky
Crystal Annealing
Flash-cooling crystals can sometimes increase the mosaicity of biological macro-
molecular crystals. In some cases, macromolecular crystal annealing can reduce
the mosaicity of flash-cooled crystals without affecting molecular structure.
Crystal annealing involves cycling a flash-cooled crystal to room temperature and
then back to cryogenic temperature. The procedure can also be applied to some-
times restore diffraction from flash-cooled crystals that were not properly han-
dled to and from cryogenic storage. Crystal annealing does not seem to improve
a poorly diffracting crystal suffering from molecular disorder. The essential
features of the crystal annealing procedure are the following: The crystal is first
flash-cooled. The crystal is removed from the cryostream and quickly transferred
to a drop of cryoprotectant at crystal growth temperature and allowed to remain
in the drop for at least three minutes. The drop should be covered to prevent
evaporation. Finally, the crystal is remounted on a loop and flash-cooled.
Reference:
Macromolecular Crystal Annealing: Overcoming Increase Mosaicity Associated with Cryocrystallography, J.M. Harp, D.E.
Timm, and G.J. Bunick, Acta Cryst. (1998) D54 622-628.
Halides for Phasing Proteins
In the publication, Novel approach to phasing proteins: derivatization by short
cryo-soaking with halides, Dauter et al. described the procedure with four
different proteins including lysozyme, RNAase A, subtilisin, and xylanase. Hey!
Congratulations for taking the time to test beyond lysozyme! We wish more folks
would take the time to test novel crystallization discoveries with more than just
lysozyme! Anyway, Sodium iodide or Sodium bromide was added to the crystal-
lization reagent and cryoprotectant mixture for the derivatization. It was noted
that 1.0 M Sodium bromide did not seem to affect the quality and diffracting
power of the tested crystals. 1.0 M Potassium iodide did mess with the crystal-
lization of lysozyme and xylanase so the Potassium iodide concentration was
reduced to 0.35 and 0.5 M for these two proteins (CrystalNews Editor Note: Both
lysozyme and xylanase were crystallized in high salt where the other two proteins
were crystallized in MPD and PEG. So it seems one might need to adjust the
NaBr or KI concentration for high salt crystallizations and perhaps not too much
for polymer or non-volatile organic crystallizations. Who knows? More results
needed for a consensus). Crystals were transferred to the halide laced mother
liquor for 15 to 45 seconds. Crystals were then flash-cooled (you are welcome
Sean) for data collection. A fifth protein, in fact, a new protein to be described in
a future publication was also successfully tested with this novel protocol.
For more details about the identification of anamolous sites, the phasing, and
application to an unknown structure, see the following reference about quick
cryo-soaking with halides as a possible alternative method for phasing protein
crystal structures.
Reference:
Acta Cryst. 2000, D56, 232-237.
1991-2009 Hampton Research Corp. All rights reserved. Printed in the United States of America.
This guide or parts thereof may not be reproduced in any form without the written permission of the publishers.
how to:
cryocrystallography
and the crystalcap system
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Solutions for Crystal Growth
Mounting Crystal
With the CrystalCap and
CryoLoop on the end of the
CrystalWand, remove a
crystal from the drop.
Place Crystal in Cryostream
Place the CrystalCap onto
the magnetic platform on the
goniometer head properly posi-
tioned in the cryostream.
Using Dewar to Freeze Crystal
Use the CrystalWand to immerse the
CrystalCap and CryoLoop containing the
crystal into a dewar containing liquid
nitrogen to freeze the crystal.
Storing Frozen Crystal
Immerse the CryoTong contain the
CrystalCap in a dewar contain-
ing liquid nitrogen. Place the
CrystalCap onto the vial and store
the crystal.
Use the Vial Clamp to hold the
CrystalCap vial and position the
CrystalCap into the vial.
Place the vial in a cryocane and
store the cryocane in a liquid
nitrogen storage dewar.
Using the CryoTong
Immerse the CryoTong in
liquid nitrogen and place
the CrystalCap into the
CryoTong.
Transferring the Crystal
Move the CryoTong from the
dewar and position the CrystalCap
onto the magnetic platform on the
goniometer head so the crystal is
positioned in the cryostream.
Removing Crystal and Storing
Immerse the CryoTong in liquid nitrogen. Place
the CryoTong over the CrystalCap and crystal.
Tilt and remove the CrystalCap from the goni-
ometer head.
Direct Freezing
Dewar Freezing
Storing Crystal
Transferring Crystal
Store Crystal
f l o w c h a r t f o r c r y o c o o l i n g o f c r y s t a l s
1991-2009 Hampton Research Corp. All rights reserved. Printed in the United States of America.
This guide or parts thereof may not be reproduced in any form without the written permission of the publishers.
Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com 204 Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com
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usi ng hali des for phasi ng
molecular st ructures
Solutions for Crystal Growth
Preliminary research shows that bromide (as well as iodide) can diffuse into protein crystals when soaked with the appropriate solution and can successfully be
used for phasing.
1
Halide soaked crystals can be used for MAD, as well as multiple or single isomorphous replacement with anomalous scattering or for single
anomalous diffraction. The procedure has been termed Halide Cryosoaking by Dauter and Dauter.
1
In simplest terms, the procedure involves dipping the crystal
for a short period of time into a cryoprotectant solution that contains a significant concentration of halide salt. Although no single recipe will suffice for all proteins
since each crystal has a unique crystallization recipe and will require different cryoprotectant cocktails, there are some general suggestions to follow at this time.
First, there are currently more successful examples using bromide than iodide. The soak time is approximately 10 to 20 seconds. Longer soaks have not lead to
more incorporation of halide ions for the examples to date. The concentration range of sodium bromide for soaking is approximately 0.25 to 1 M. Higher concen-
trations of halide ions may lead to more sites with higher occupancies and increased phasing power. Factors influencing the success of the procedure include the
resolution and quality of the x-ray diffraction data, crystal symmetry, packing density, and pseudo-symmetric arrangements of molecules.
Tips for successful Halide cryosoak include:
1. Initially, preserve the formulation of the crystallization reagent used to grow the crystal as well as the formulation for a successful cryosoak and then
add the halide salt. In other words, leave everything constant and add the halide salt.
2. If the crystallization reagent contains salt, try substituting the halide salt, especially if the salt is Sodium chloride.
3. High concentrations of the halide salt can serve as a cryoprotectant without the addition of other traditional cryoprotectants (Glycerol, MPD, sucrose).
4. Experiment with soak conditions. Vary the concentration of the original reagents, the concentration of the halide salt, and the soak time.
Examples of successful crystallization reagents optimized for halide cryosoaking:
Original Condition (Black) Halide Cryosoak Condition (Blue)
1.0 M Ammonium sulfate, 5 mM guanidine, 10% Glycerol, 0.1 M Sodium citrate pH 3.3
3
1.0 M Ammonium sulfate, 5 mM guanidine, 18% Glycerol, 0.1 M Sodium citrate pH 3.3, 1 M Sodium bromide
3
1.4 M Lithium sulfate, 0.1 M Tris pH 7.5
3
1.2 M Lithium sulfate, 0.1 M Tris pH 7.5, 1 M Sodium bromide, 14% Glycerol
3
1.0 M Sodium chloride, 0.1 M Sodium acetate pH 4.7
2
0.1 M Sodium acetate pH 4.7, 1.0 M Sodium bromide, 30% Glycerol
2
50% MPD, 0.1 M Sodium acetate pH 5.4
2
50% MPD, 0.1 M Sodium acetate pH 5.4, 1.0 M Sodium bromide
2
12% PEG 4,000, 0.1 M Citrate, 1.0 M Sodium chloride, 10 mM Calcium chloride, pH 6.0
2
12% PEG 4,000, 10 mM Citrate, 10 mM Calcium chloride, 25% Glycerol, 1.0 M Sodium bromide
2
10% Ammonium sulfate, 0.1 M TRIS HCl pH 7.4
2
10% Ammonium sulfate, 0.1 M TRIS HCl pH 7.4, 25% Glycerol, 1.0 M Sodium bromide
2
References
1. Entering a new phase: Using solvent halide ions in protein structure determination. Dauter, Z. & Dauter, M., Structure, Vol 9, R21-26, Feb 2001.
2. Novel approach to phasing proteins: derivatization by short cryo-soaking with halides. Dauter, Z., Dauter, M., & Rajashankar, K.R. Acta Cryst. (2000) D56, 232-237.
3. Practical experience with the use of halides for phasing macromolecular structures: a powerful tool for structural genomics. Dauter, Z., Li, M. and Wlodawer, A.
1991-2009 Hampton Research Corp. All rights reserved. Printed in the United States of America.
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205
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Solutions for Crystal Growth
mi croseedi ng
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Background
A crystallization experiment typically begins with the sample in a stabilizing
solution of water and a buffer, salt, reducing agent, ligand, or other reagent.
Prior to mixing the sample with crystallization reagent, this sample solution is
undersaturated with respect to the macromolecule in question (sample). In an
undersaturated sample solution, no crystals can nucleate, nor can crystals grow
from seeds. Upon addition of a crystallization reagent, the relative supersatura-
tion of the sample is increased. Assuming the crystallization reagent decreases
the solubility of the sample to increase the relative supersaturation, three events
can take place. In the first stage of supersaturation, the metastable zone, spon-
taneous homogenous nucleation cannot occur, but crystal growth from seeds
can occur. Moving further into supersaturation, the labile zone, spontaneous
homogeneous nucleation and crystal growth can occur. Further into supersatu-
ration, the precipitation zone, precipitation of the sample from solution occurs.
See figure 1 below.
Seeding
Seeding allows one to grow crystals in the metastable zone. Why would one
want to do this? For control, reproducibility, and to improve the likelihood of a
successful crystallization experiment. In the metastable zone, crystals can grow
from seeds but cannot spontaneously nucleate. By placing a seed or solution of
seeds in a drop which is saturated to this zone, one can use the seeds to grow
larger single crystals. By controlling the number of seeds introduced into the
zone drop, one can control the number of crystals grown. It is not practically
possible to measure and know the number of seeds introduced to a drop, but
by performing serial dilutions from a concentrated seed stock, one can control
the number of crystals grown in the drop.
Seeding Examples
Example 1: (A) Initial crystals of the catalytic domain of CheB methyltransferase
were grown from Crystal Screen 2, reagent 42. (B) Microseeds were taken
from (A) to grow diffraction quality crystals.
Example 2: (A) Initial crystals of CheR methyltransferase were grown producing
platelet clustered crystals. (B) Microseeds were taken from (A) to grow quality
3-dimensional crystals.
Thank you Ann West and Snezana Djordjevic for your examples of microseed-
ing.
example 1
example 2
A
B
A
B
Stable
(undersaturated)
Metastable
(supersaturated)
Labile
(supersaturated)
Salt Concentration
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Precipitation
figure 1
The diagram is divided into 4 zones:
1. Stable: undersaturated where crystallization
is not possible.
2. Metastable: supersaturated where nuclei
cannot form but crystals from seeds can grow.
3. Labile: supersaturated where crystallization
can occur.
4. Precipitation
1991-2009 Hampton Research Corp. All rights reserved. Printed in the United States of America.
This guide or parts thereof may not be reproduced in any form without the written permission of the publishers.
Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com 206 Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com
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Solutions for Crystal Growth
det ergent s at work
The image below is a view of the crystal packing of -lactamase. The image
demonstrates the detergent molecule linking SHV-1 molecules in the crystal
lattice. The detergent binding to the -lactamase is CYMAL

-6. It appears as
though the well-ordered detergent molecules are linking protein molecules
in the crystal lattice. The hydrophobic tail of the detergent is buried in the
protein and the hydrophilic head is hydrogen bonded with a neighboring
molecule.
-lactamase resisted producing any decent crystals until the detergents CYMAL

-5
and CYMAL

-6 were included in the crystallization drop. The crystals were grown

in a relatively low ionic strength environment (15% w/v PEG 6,000, 50 mM HEPES
pH 7.0, and detergent) via sitting drop vapor diffusion.
The crystals and the structure demonstrate the efficacy of detergents as additives to
improve crystallization of soluble proteins and bring to light the molecular linking
capabilities that small molecules can have in biological macromolecular crystals.
CYMAL

-5 and CYMAL

-6 are part of the Hampton Research Detergent Screen.

Images and discussion courtesy of James Knox, University of Connecticut,
Department of Molecular and Cell Biology 1999.
Submitted by Ivana Kuta Smatanova, University of South Bohemia, Photosynthesis
Research Center and Laboratory of Biomembranes, Branisovska 31, CZ-370 05
Ceske Budejovice, Czech Republic
Crystallization trials have been performed in Cryschem Plates for sitting drops
and in capillary tubes at 277K or 289K. 15-mg/ml (1.3 mg/ml chlorophyll a)
protein has been mixed with different type of detergents and additives used
in the crystallization experiments. We have either used the Hampton Research
MembFac kit for membrane protein screening or solutions prepared in our
own lab. 50 commercial solutions and 50 different kind of detergents have
been available for experiments. According to the published information about
a crystallization of photosynthetic reaction center core complexes we have used
solutions containing PEG as precipitant. Concerning detergents we have pro-
ceeded in accordance with literature data and up to this day we have tried 15
different detergents, detergents with different alkyl chain length and mixtures
of detergent as well. Processes inside the drops have been observed during the
period of 2-4 weeks at 289K and 4-6 weeks at 277K. The major part of drops
contained precipitates, separated phases or segregated carotenoids in various
forms. We have already found for crystallization experiments acceptable pH rate
around 7.00 (0.50), the precipitant PEG 4000 or PEG 6000 (in the case of using
a commercial solutions without PEGs, we have observed the clear drops without
precipitate or other formations). For detergents, in the presence of 1-heptyl--
D-glucoside, 1-octyl--D-glucoside, 1-nonyl--D-glucoside and their thio forms
such as heptyl--D-thioglucoside and 1-octyl--D-thioglucoside, different type
of carotenoid formations are separated from the protein samples. We have also
performed crystallization experiments either in the absence or in the presence
of additional amphiphiles (heptanetriol) that has been used to modify detergent
characteristics and to facilitate membrane protein crystallization.
1
Nowadays crys-
tallization experiments are still in the progress; we have tried to use new kind of
detergents and their mixtures and new crystallization conditions as well.
Reference
1. Marone P.A., Thiyagarajan P., Wagner A.M. and Tiede, D.M. Effect of detergent alkyl chain length on crystallization of a
detergent-solubilized membrane protein: correlation of protein-detergent particle size and particle-particle interaction with
crystallization of the photosynthesis reaction center from Rh. sphaeroides J. Cryst. Growth 207 (1999) 214-225.
c ry s t a l l i z a t i o n e x p e r i me nt s o n t he p ho t o s y s t e m I I
r e a c t i o n c e nt e r f r o m p i s um s a t i v um
1991-2009 Hampton Research Corp. All rights reserved. Printed in the United States of America.
This guide or parts thereof may not be reproduced in any form without the written permission of the publishers.
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P A G E 1 - 2
Solutions for Crystal Growth
The following crystallization recipes can be useful for teaching and practicing
protein crystallization or for fooling your PI into thinking you have grown huge
crystals of that impossible-to-crystallize protein.

Lysozyme
Protein source: Hampton Research HR7-108 or HR7-110
Worthington Biochemical Corporation
Sigma-Aldrich
Recommended concentration: 20 to 100 mg/ml
Sample Buffer: 0.1 M Sodium acetate pH 4.6
Crystallization reagent: 8% w/v Sodium chloride, 0.1 M Sodium acetate pH 4.6
10% w/v Sodium chloride, 0.1 M Sodium acetate pH 4.6, 10% v/v Glycerol
Additional Reagents: Hampton Research Index screen reagents 8, 22, 28, 31,
34, 40, 58, 59, 69, 86, 88 and Hampton Research Grid Screen Sodium Chloride
reagents
Method: Hanging or sitting drop vapor diffusion, microbatch, free interface
diffusion
Procedure: Mix equal amounts of lysozyme with reagent, incubate at 4C or room
temperature
Lysozyme crystals in 15 minutes
Recommended concentration: 100 mg/ml
Sample Buffer: 0.1 M Sodium acetate pH 4.6
Crystallization reagent: 30% w/v PEG MME 5,000, 1.0 M Sodium chloride, 0.05 M
Sodium acetate trihydrate pH 4.6 (Hampton Research HR2-805)
Method: Hanging or sitting drop vapor diffusion, microbatch, free interface
diffusion
Procedure: Mix equal amounts of lysozyme with reagent, incubate at 4C or room
temperature

Thaumatin
Protein source: Worthington Biochemical Corporation or Sigma-Aldrich
Recommended concentration: 20 to 40 mg/ml
Sample buffer: DI water, 25 mM BIS-TRIS propane or Imidazole pH 6.5
Crystallization reagent: 1.0 M Potassium sodium tartrate
Additional reagents: Hampton Research Index screen reagents 26, 31, 78, 86
Method: Hanging or sitting drop vapor diffusion, microbatch, free interface
diffusion
Procedure: Mix equal amounts of protein with reagent, incubate at 4C or room
temperature

Glucose isomerase
Protein Source: Hampton Research HR7-100 or HR7-102
Recommended concentration: 10 to 40 mg/ml
Sample buffer: 10 mM HEPES, 1 mM Magnesium chloride hexahydrate
Crystallization reagent:
1.5 - 2.5 M Ammonium sulfate pH 6.0 - 9.0
10 - 15 % w/v PEG 4,000 - 8,000, 0.2 M salt *, 6.0 - 9.0
0.6 - 1.0 M Sodium citrate tribasic dihydrate, pH 6.0 - 8.0
0.1 - 0.3 M Magnesium formate dihydrate
1.0 M Sodium formate, pH 5.0 - 7.0
10 - 20 % v/v MPD, 0.2 M salt *, pH 6.0 - 9.0
10 - 20 % v/v PEG 400, 0.2 M salt *, pH 6.0 - 9.0
10 - 20 % 2-Propanol, 0.2 M salt *, pH 6.0 - 9.0
* Ammonium sulfate, Magnesium acetate, Magnesium chloride, Calcium acetate
and other salts
Additional reagents: Hampton Research Index screen reagents including but not
limited to 4, 5, 14, 48, 49, 67, 92
Method: Hanging or sitting drop vapor diffusion, microbatch, free interface
diffusion
Procedure: Mix equal amounts of protein with reagent, incubate at 4C
Glucose isomerase for cryo
Special thanks to Laurie Betts for the the cryo work and data!
Protein Source: Hampton Research HR7-100 or HR7-102
Recommended concentration 20 to 30 mg/ml
Sample buffer 10 mM HEPES, 1 mM Magnesium chloride hexahydrate
Crystallization reagent: 30% v/v 2-Methyl-2,4-pentanediol, 0.1 M Sodium
Cacodylate pH 6.5, 0.2 M Magnesium Acetate tetrahydrate (Crystal Screen Cryo
reagent 21) or 30% v/v Polyethylene Glycol 400, 0.1 M HEPES - Na pH 7.5, 0.2 M
Magnesium Chloride hexahydrate (Crystal Screen Cryo reagent 23)
Method: Hanging or sitting drop vapor diffusion, microbatch, free interface
diffusion
Method: Mix equal amounts of Glucose isomerase and reagent. Mount crystal
in CryoLoop. Swish mounted crystal through reservoir to increase MPD or PEG
400 concentration in crystal to decrease mosaicity. No swish for more mosaicity.
Crystal can diffract to 1.6 Angstrom, indexes in I-centered orthorhombic with
a=92.7 b=97.4 c=102.7. Mosaicity is about 0.5 to 0.6.

Alpha Lactalbumin
Protein source: Sigma-Aldrich
Recommended concentration 10 to 20 mg/ml
Sample buffer: 10 mM TRIS hydrochloride pH 8.5
Crystallization reagent: 30% w/v Polyethylene glycol 3,350, 0.1 M TRIS hydrochlo-
ride pH 8.5, 0.2 M Lithium sulfate
Additional reagents: Hampton Research Index screen reagent including but not
limited to
4, 5, 6, 74
Method: Hanging or sitting drop vapor diffusion, microbatch, free interface
diffusion
Procedure: Mix equal amounts of protein with reagent, incubate at 4C or room
temperature

Catalase
Protein source: Worthington Biochemical Corporation or Sigma-Aldrich
Recommended concentration: 10 to 20 mg/ml
Sample buffer: 10 mM HEPES pH 7.0
Crystallization reagent: 20% w/v Polyethylene glycol 3,350, 0.1 M HEPES pH 7.0
Additional reagents: Hampton Research Index screen reagents including but not
limited to 39, 55, 63, 64, 88, 89
Method: Hanging or sitting drop vapor diffusion, microbatch, free interface
diffusion
Procedure: Mix equal amounts of protein with reagent, incubate at 4C or room
temperature
protei n crystalli zati on
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Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com 208 Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com
Solutions for Crystal Growth
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P A G E 2 - 2
1991-2009 Hampton Research Corp. All rights reserved. Printed in the United States of America.
This guide or parts thereof may not be reproduced in any form without the written permission of the publishers.
Ribonuclease S
Protein source: Sigma-Aldrich
Recommended concentration: 40 mg/m
Sample buffer: 10 mM HEPES pH 7.0
Crystallization reagent: 30% Polyethylene glycol 4,000, 0.1 M Sodium citrate
tribasic dihydrate pH 5.6, 0.2 M Ammonium acetate
Additional reagents: Hampton Research Crystal Screen reagent including but not
limited to 9, 20
Method: Hanging or sitting drop vapor diffusion, microbatch, free interface dif-
fusion
Procedure: Mix equal amounts of protein with reagent, incubate at 4C or room
temperature
Proteinase K
Protein source: Sigma-Aldrich
Recommended concentration: 20 mg/ml
Sample buffer: 10 mM HEPES pH 7.0
Crystallization reagent: 0.4 M Potassium sodium tartrate tetrahydrate
Additional reagents: Hampton Research Crystal Screen reagent including but not
limited to 2, 3, 4, 6, 7, 9, 10, 11, 15, 16, 17, 18, 20, 22, 25, 28, 29, 30, 31, 32, 33, 34,
35, 36, 38, 39, 40, 41, 42, 44, 45, 46, 47, 48, 49, 50
Method: Hanging or sitting drop vapor diffusion, microbatch, free interface dif-
fusion
Procedure: Mix equal amounts of protein with reagent, incubate at 4C or room
temperature
Trypsin (bovine pancreas)
Protein source: Worthington Biochemical Corporation or Sigma-Aldrich
Recommended concentration: 15 to 60 mg/ml
Sample buffer: 10 mM Calcium chloride, 10 mg/ml Benzamidine hydrochloride,
20 mM HEPES pH 7.0
Crystallization reagent: 4% w/v Polyethylene glycol 4,000, 0.2 M Lithium sulfate
monohydrate, 0.1 M HEPES pH 7.0, 15% v/v Ethylene glycol
Additional reagents: Hampton Research Crystal Screen reagents including but
not limited to 4, 15, 16, 20, 28, 30, 31
Method: Hanging or sitting drop vapor diffusion, microbatch, free interface dif-
fusion
Procedure: Mix equal amounts of protein with reagent, incubate at 4C or room
temperature
Ferritin
Protein source: Sigma-Aldrich
Recommended concentration: 20 mg/ml
Sample buffer: 10 mM HEPES pH 7.0
Crystallization reagent: 0.8 Ms Ammonium sulfate, 0.1 M HEPES pH 7.5, 60 mM
Cadmium sulfate
Additional reagents: Crystal Screen 2 reagents 12, 34 and Index screen reagent 64
and PEG/Ion 2 reagent 42 and PEGRx 2 reagent 20
Method: Hanging or sitting drop vapor diffusion, microbatch, free interface dif-
fusion
Procedure: Mix equal amounts of protein with reagent, incubate at 4C or room
temperature
Xylanase
Protein source: Hampton Research HR7-104, HR7-106
Recommended concentration: 3 to 20 mg/ml
Sample buffer: 0.09 M Sodium potassium phosphate pH 7, 21% v/v Glycerol
Crystallization reagent: 1.0 M Sodium potassium phosphate pH 7
Additional reagents: Quik Screen reagents A5, B5, C5, D5, A6, B6, C6, D6
Method: Hanging or sitting drop vapor diffusion, microbatch, free interface dif-
fusion
Procedure: Mix equal amounts of protein with reagent, incubate at 4C or room
temperature
Lipase B
Protein source: Hampton Research HR7-099
Recommended concentration: 10 to 100 mg/ml
Sample buffer: Deionized water or 10 mM Sodium acetate pH 4.6
Crystallization reagent: 1.0 M Sodium potassium phosphate pH 7
Additional reagents: Quik Screen reagents A5, B5, C5, D5, A6, B6, C6, D6
Method: Hanging or sitting drop vapor diffusion, microbatch, free interface dif-
fusion
Procedure: Mix equal amounts of protein with reagent, incubate at 4C or room
temperature
Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com
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P A G E 1 - 2
Solutions for Crystal Growth
temperature as a
crystalli zati on vari able
Temperature and Crystallization
Temperature can be a significant variable in biological macromolecule and small
molecule crystallization.
(1-5)
Temperature often influences nucleation and crys-
tal growth by manipulating the solubility and supersaturation of the sample.
Temperature has also been shown to be an important variable with phase separa-
tion in detergent solutions during membrane protein crystallization.
7
Control and manipulation of temperature during the screening, optimization and
production of crystals is a prerequisite for successful and reproducible crystal
growth of proteins with temperature dependent solubility. Christopher et al.,
5
testing 30 randomly chosen proteins, found 86% demonstrated a temperature
dependent solubility and suggested that temperature induced crystallization
could be a generally useful technique. Temperature was shown to affect quan-
tity, size, and quality of the crystals, as well as sample solubility and preliminary
crystallization data.
One advantage of temperature is that it provides precise, quick, and reversible
control of relative supersaturation. Using temperature, in addition to standard
crystallization variables such as pH, reagent composition and concentration,
and sample concentration, can increase the probability of producing crystals as
well as uncover new crystallization conditions for a sample. Additional crystal-
lization conditions may uncover reagent formulations more amicable to heavy
atom derivatization, cryoprotection, and optimization or at least offer options.
Temperature is amenable to control and can be used to carefully manipulate
crystal nucleation and growth. This control can also be used to etch or partially
dissolve the crystal and then grow it back in an attempt to improve size, mor-
phology, and quality or assist with seeding. Temperature control is noninvasive
and can manipulate sample solubility and crystallization with altering reagent
formulation.
Typically, crystallization screens and experiments are performed at room tem-
perature and possibly 4C. A reasonable range of temperature to screen and
optimize for protein crystallization is 4 to 37C and some proteins have been
crystallized at 60C (glucagon and choriomammotropin). Temperature incuba-
tions above room temperature should be monitored closely for evaporation from
the drop and reservoir. A 2 l hanging drop vapor diffusion experiment at 37 C
can evaporate in as little as 48 hours depending upon the plate and the quality
of seal. Microbatch under Paraffin Oil can minimize evaporation problems. In
the case of room temperature incubations, temperature control and stability are
often minimal since the experiments may be left in the open room. In an open
room, temperature fluctuations may be significant, especially over a 24 hour
period and on weekends when thermostatic control of the room environment
can fluctuate 10 degrees or more. Incubation at 4C and other temperatures are
often more stable since the incubation is performed in some type of incubator.
Another source of temperature fluctuation occurs while viewing experiments.
The light microscope is a heat source and extended viewing can significantly alter
the temperature of small drops. Quick, efficient viewing can minimize tempera-
ture changes. Also, remember to turn off the light source when leaving plates on
the stage in one position for more than a few seconds.
While controlled temperature can be important for consistent results, tempera-
ture fluctuation can be useful in obtaining high quality crystals by screening a
larger range of crystallization conditions.
8
For a sample with temperature depen-
dent solubility, changes in temperature can equate to changes in a crystallization
reagent condition. Hence, a sparse matrix screen takes on a new dimension when
screened at multiple temperatures, or ramped over several different tempera-
tures over a period of time.
How does one test for the effect of temperature and temperature dependent
solubility without consuming a lot of sample? One solution is to set a single
crystallization screen at one temperature, allow the experiment to incubate for a
week, record the results, and then move the plate to another temperature. Allow
the experiment to incubate for a few days to a week at the new temperature and
record the results. If one notices changes is solubility (i.e. clear drop turning to
precipitate, or precipitate turning to clear drops) between the two temperatures,
then the sample has temperature dependent solubility and temperature should
be explored as a crystallization variable.
Temperature gradients can be used for screening and optimization of proteins
with temperature dependent solubility. For screening, set the experiment at one
temperature, allow it to equilibrate, and then slowly change the temperature to
a second temperature. In general, ramp the temperature so that the sample is
exposed to an increase in relative supersaturation as the temperature changes
over time. In other words, ramp from high to low temperature if the sample is
more soluble at high temperatures. This can be accomplished using a program-
mable temperature incubator. A temperature gradient or ramp, allows one to
slowly approach temperatures where a sample may have a decrease in solubility
with a corresponding increase in relative supersaturation. Published examples of
temperature gradient or temperature ramp crystallization include elastase9 (25
to 20C gradient), alpha-amylase10 (25 to 12C gradient), and insulin11 (50 to
25C gradient).
To demonstrate how screening temperature could affect and enhance the results
obtained from a preliminary crystallization screen, a temperature incubator was
used to screen four different temperatures. Using Glucose isomerase and Crystal
Screen, sitting drop vapor diffusion experiments were set using Cryschem
plates at 4, 15, 25, and 37C. Drops were observed daily and the results were
quite interesting. Glucose Isomerase crystallized in 19 conditions at 25C, 23
conditions at 15C, 28 conditions at 4C, and 12 conditions at 37C. A similar
approach with Trypsin, yielded crystals in 8 conditions at 15C, 4 conditions at
25C, and 7 conditions at 32C. In the case of Trypsin, a single set of Cryschem
plates were set and then simply moved from one temperature to another over a
period of one week, scoring results before each temperature change.
Temperature Tips
For proteins with "normal" solubility, in high salt they will be more soluble at
cold than at warm temperatures.
For proteins with "normal" solubility, in low salt they will be more soluble at
warm than at cold temperatures.
continued next page....
Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com 210 Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com
Solutions for Crystal Growth
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temperat ure as a
cryst alli zat i on vari able
P A G E 2 - 2
1991-2009 Hampton Research Corp. All rights reserved. Printed in the United States of America.
This guide or parts thereof may not be reproduced in any form without the written permission of the publishers.
For proteins with "normal" solubility, they will precipitate or crystallize from
lower concentration of PEG, MPD, or organic solvent more slowly at low
than at high temperatures.
Temperature effects can be more pronounced in low ionic strength reagent
conditions.
Do not use the appearance or non-appearance of crystals at various tempera-
tures to gauge the effectiveness of temperature as a crystallization variable.
Rather, use the difference in the solubility at different temperatures to gauge
the effect temperature has on sample solubility. If an effect is observed,
explore temperature as a crystallization variable.
Temperature can affect different crystal forms and growth mechanisms.
12
When incubating experiments below and above room temperature and view-
ing experiments at room temperature, condensation can be a problem. To
minimize and avoid condensation with vapor diffusion experiments, stack a
dummy plate (with the reservoir flled with water and sealed) at the bottom
and top of the stack of plates. This will slow the temperature change in the
sandwiched plates and minimize condensation.
The microbatch method works well for temperature exploration. In a
traditional microbatch experiment, the relative supersaturation of the system
does not change since in theory, there is no vapor diffusion. However, if the
sample exhibits temperature dependent solubility, temperature can be used
to manipulate sample solubility in a microbatch experiment. Another plus of
using microbatch is the lack of condensation while viewing the experiment.
Covers of microbatch plates can be removed for a clear view.
Condensation with a hanging drop can lead to an alteration when the con-
densation mixes with the drop. In a sitting drop, condensation does not mix
with the drop unless the condensation falls from above.
To dry up condensation, add a small amount of concentrated salt solution to
the reservoir. Keep in mind this might also further dehydrate your drop.
Ideally, one should set the experiment at the eventual incubation tempera-
ture and all reagents, samples, and plates should be equilibrated to the in-
cubation temperature. This is a reality for room temperature and 4C setups
for those of us with cold rooms. For the rest of us, we can set the experi-
ment at room temp and then toss it into the incubator. Or, for 4C setups,
one can cheat. Simply incubate the reagents, sample, plates and slides in the
refrigerator before set up. During the set up, place materials in a tray full of
ice. Maintain the plates on ice during the set up. Seal and move smartly to
the 4C incubator.
Nucleic acid temperature stability allows one to examine temperatures
between 4 and 35C.
Increasing temperature increases the disorder of reagent molecules. Varying
the temperature of a crystallization experiment can manipulate sample-
sample as well as sample-reagent and reagent-reagent interactions. Such
manipulations may have an impact on interactions which control nucleation
and crystal growth. In addition, such interactions may have an impact on
crystal packing as well as the termination of crystal growth.
Temperature can be a habit modifer and change the crystal lattice. For ex-
ample, at temperatures below 25C and in the presence of sodium chloride
and acidic pH, the tetragonal form of lysozyme is favored. Under similar
reagent conditions above 25C, the orthorhombic form is favored.
13
The preparation of heavy atom isomorphous derivatives can depend upon
the temperature of the experiment. In most cases, it seems the soak tem-
perature is the same as the crystallization temperature.
References
1. Giege, R., and Mikol, V., Trends in Biotechnology (1989) 7, 277.
2. McPherson, A., European J. Biochemistry (1990) 189, 1.
3. A. Ducruix and R. Giege, Editors, Crystallization of Nucleic Acids and Proteins: A Practical Approach, IRL Press at Oxford
University Press, 1991.
4. Lorber, B., and Giege, R., Journal of Crystal Growth (1992) 122, 168-175.
5. Christopher, G.K., Phipps, A.G., and Gray, R.J., Journal of Crystal Growth (1998) 191, 820-826.
6. Haser, R., et al., Journal of Crystal Growth (1992) 123, 109-120.
7. Garavito, R.M., and Picot, D., Journal of Crystal Growth (1991) 110, 89.
8. Drenth, J., Crystal Growth (1988) 90, 368.
9. Shotton, D.M., Hartley, B.S., Camerman, H., Hofmann, T., Nyborg, S.C., and Rao, L., Journal of Molecular Biology (1968)
32, 155-156.
10. McPherson, A., and Rich, A., Biochem. Biophys. Acta (1972) 285, 493-497.
11. T.L. Blundell, and L.N. Johnson, Protein Crystallography, Academic Press (New York) 1976, 59-82 (method by Guy
Dodson).
12. A. McPherson, Crystallization of Biological Macromolecules, Cold Spring Harbor Laboratory Press, 1999.
13. Ataka, M., and Tanaka, S., Biopolymers (1986) 25, 337.
Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com
Crystal image.
Anastasia Tziridis, Institute for Biotechnologie
Martin-Luther-Universitt Halle-Wittenberg, Germany
Crystals of human mineralocorticoid receptor complexed
with ligand. Grown using Hampton Salt Rx screen.
Kevin P Madauss, GlaxoSmithklinem Research Triangle Park,
North Carolina, USA.
Protein crystal grown by counter diffusion technique in
0.2mm capillary in the presence of 0.1 % agarose.
Pavlina Rezacova,
UT Southwestern Medical Center at Dallas, Texas, USA.
ti ps from ramc
Crystals of the Deinococcus radiodurans 50S ribosome subunit.
From the group of Paola Fucini, Max Planck Institute for Molecular Genetics, Berlin, Germany.
213
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Filtering the Protein
Naomi Chayen
To reduce the number of crystals and increase their size, try filtering the protein
solution prior to setting up the experiment. Try the following filter sizes: 0.22,
0.1 micron and 300 kD molecular weight cut-off. Try the Millipore centrifugal
filters.
Pseudo-Microseeding
Mike Sintchak
When working with crystals that grow fairly rapidly (one day) try the following.
Pipette multiple protein drops (2 to 4 works best) onto the cover slide. Using
a single pipette tip, get the reservoir solution to mix with the first drop. Now,
go back into the reservoir with the same tip to get the reservoir solution for the
second drop. Continue for the remaining drops with the same tip. In certain
cases, seeding starts very quickly, so by using the same tip one can introduce
minute seeds to successive drops. Use the same cover slide with multiple drops
to minimize evaporation.
Low Molecular Weight PEGs
Lesley Haire
When screening with low molecular weight PEGs try microbatch. Crystals appear
rapidly with PEG 400-2000. To convert to vapor diffusion use 0.2 M buffer in the
well and a 1:1 drop ratio. Try using a positive displacement pipette such as the
Anachem Microman 1 - 10 l. These are much more accurate.
Purest is Not the Best
Michal Harel
A protein which was purified and showed some faint bands of contaminants on a
native gel was crystallized and solved successfully. The same protein, purified by
HPLC and resulting in a single band native gel did not crystallize.
Cryoprotectant
Elspeth Garman
When making up cryoprotectant solutions containing glycerol, put a test tube of
glycerol in a beaker of warm water. The viscosity falls and it is easier to pipette
accurately.
Mass spectrometry
David Leys
We found mass spectrometry like ESMS and MALDI highly efficient in determin-
ing impurities and/or microheterogeneities in our protein sample/batch. In most
cases it is a simple, straight forward method which requires a minimum amount
of sample. In some cases it has shown to detect impurities/microheterogeneities
when other techniques did not.
Shape of Drop
Ursula Kamlott, Hoffmann-La Roche: ursula.kammlott@roche.com
One of my proteins produced only zillions of tiny useless crystals when I mixed
the drops the conventional way-mixing well, overlaying, etc. the protein with
precipitant solution. Large, gorgeous crystals were produced when I crossed the
drop, creating a gradient within the drop. This worked best, setting up sitting
drops with vapor diffusion.
Dont Flip
Dennise Dombroski
When removing crystals from a hanging drop, I sometimes find that the biggest
crystals fall against the coverslip and are impossible to resuspend without dam-
age. I had our glass shop make a stand to transfer the coverslip that enabled me
to manipulate the crystals more easily.
Extreme Soak
Jirundon Yuvaniyama
For soaking crystals with compounds with limited solubility I have tried two
extreme ways (although not much- and more experiments should be tried):
Leave some solid compound in the soaking solution.
Dissolve some compound in n-octanol and layer the octanol solution on top
of the soaking experiment.
These two provide (hopefully) more or less constant concentration of the com-
pound in the soaking solution. The octanol layer may help reduce air oxidation
by preventing direct contact of soaking solution to air.
I found that 200 l of soaking solution in a well of the 24 well Linbro

plate is a
good volume to work with:
Not too little that possibly causes concentration change due to evaporation
(Sealed well) and not too much that the crystals get lost in the solution.
The Glycerol Effect
Anil Mistry
Glycerol has many benefits but also some drawbacks. We found it to be beneficial
with one protein we were working with; this protein is a transpeptidase called Mur
A. The protein is quite soluble and could concentrate to 20 mg/ml but it lost activi-
ty over time when stored at 80C. We therefore dialysed it into 50% v/v Glycerol to
see if the activity could be retained for longer, this would allow us to make a large
batch rather than regular smaller batches, for crystallisation. On dialysis we found
a significant reduction in the volume of protein, so much so that the protein had
concentrated from 5-10 mg/ml to almost 50 mg/ml. Activity was also found to be
retained with no significant loss after 6 months at 80C. This gave us a method for
storing large batches of Mur A at 80C, without losing activity and also resulted
in a sample pre-concentrated for crystallisation and containing a cryo-protectant.
Other sugars gave similar effects; sucrose, sorbitol, etc., but none were as effective
as glycerol in achieving the 50 mg/ml final concentration.
ti ps from ramc
Tips from Recent Advances in Macromolecular Crystallization Meetings
P A G E 1 - 7
Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com 214 Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com
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Iodoacetic Acid
Bernie Santarsiero
Add a small amount of (~ 1%) iodoacetic acid to buffer solutions. This helps
prevent aggregation by carboxymethylation of cys. Also, iodoacetic acid seems
to help form salt bridges and aid in crystallization.
Soaking Crystals to Improve Resolution
Irene Weber
Try soaking poorly diffracting crystals in higher concentration of precipitate,
ammonium sulfate, or PEG. It may take several weeks so test after 1 or 2
months.
Rapid Preliminary Screening of Protein for Aggregation Using Protein
Quantities
Tom Zarembinski
When only small amounts of protein are available, it is not feasible to screen
many compounds which promote monodispersim using a dynamic light scat-
tering machine. To detect aggregation, we use a pseudo-native gel approach: 1
l of protein is mixed with 1 l of additive from Hampton Additive or Detergent
Screens. These samples are then incubated at room temperature for 20-30 min-
utes and then placed in 2x sample buffer containing no DTT, no SDS and are not
boiled. These samples are run on a standard SDS-PAGE gel. We have screened
many additives using this approach and it has given us leads for subsequent
optimization of protein buffers.
HPLC Profile
Glenn Dale
Keep an HPLC (Reverse phase) profile of your protein before crystallisation and
after crystal formation. It can be used as a quality control and tells you if any
modifications have occurred.
Dont Throw Away Without Looking Close
Kalevi Visuri
Look closely at your old test tubes when cleaning the place. Proteins do crystallize
on the walls of the tube when stored in a cold room. I picked up my tubes from the
wastebasket and an x-ray was made from an old supersaturated protein tube.
Concentration of Protein without Aggregation
Paul Reichert
Use centipreps (millipore) for concentrating protein >O.5 ml. Protein concen-
trating away from membrane. (No micro high concentration, ppt on membrane),
works very nicely for a number of proteins.
Preserve Hampton Solutions
Cheryl Janson
To preserve Hampton solution when you or a co-worker gets a hit and suspect
the Hampton solution may be magic or you cannot reproduce crystals with
lab-made solution, make the reservoir solution from lab ingredients and use
your homemade solution for reservoirs. Use the Hampton magic solution
only for the drops thus using a few l per experiment rather than O.5 ml. Saves
having a 48 well screening solution set with one tube empty and the rest at 8 ml
and still allows the superstitions or in Alexs case, the contaminated solution to
reproduce crystals.
Recycle Your Precipitate
Neali Armstrong
If your protein refolding reaction has a low yield and produces lots of precipitate
try collecting the precipitate, resolubilize in GuHCl, and refold again. This mate-
rial is sometimes more pure than the washed inclusion bodies.
90% Solutions
Anna Stevens
When optimizing or making solutions for a random scan, omit the buffer (@
final O.1 M) so that your stock is 90%. Prior to putting your stock in the well, add
100 l of 1M buffer of your choice. Next, add 900 l of 90% stock and mix. This
reduces the number of tubes for crystallization (precipitant) stocks and allows
flexibility in buffer identity and pH range. Be sure to make plenty of (~20 mL) of
precipitant, youll need 900 l per buffer.
Check Both Liquid Nitrogen + Stream Flash Cooling
Hans Parge
If your crystal does not freeze well in the cold stream try liquid nitrogen or vice
versa.
To Determine the Optimal Concentration of Your Protein for
Screen I and Screen II
Jaru Jancarik
Try (setup) drop no. 6 first (of Screen I). It should produce light precipitation if
ppt appears too heavy, reduce the protein concentration by 1/2 and try again. If
there is no precipitation in drop 6 try drop 4. If there is no precipitation in either
of the drops concentrate the protein 2 fold and try again.
Cryoprotectant Additive
Laura Pelletier
I tried adding 1-10 mg/ml BSA in the cryoprotectant when soaking crystals that
would crack. The one time it was used, the crystal did not crack and froze nicely.
I dont know if the BSA was the reason for successful freezing. I was wondering
if anyone else has tried this.
Slow Cryosoaks for Improved Crystal Stability
Bryan Prince & Melissa Harris
In a sitting drop well, cryoprotectant (20% Glycerol in crystallization buffer) should
be dribbled down the side of the well.
Crosslinking of Crystals
Clare Stevenson
When crystals are fragile or you want to transfer them to a different mother
liquor e.g. for heavy atom soaking, why not try crosslinking your crystals with
P A G E 2 - 7
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0.1% glutaraldehyde in your mother liquor. This can be done by adding the
glutaraldehyde directly to the drop or placing it next to the drop and allowing
for vapour diffusion. This crosslinking enabled us to solve Mod A at 1.2 A resolu-
tion.
Know When Enough is Enough!
Brent Segelke
As a graduate student, I spent countless hours and quantities of protein trying
to get crystals of a single construct. We never got crystals. Another group got
the structure of a homologous protein that was auto-digestive. Had we stopped
after 400 trials and altered our construct, perhaps we would have faired better,
we couldnt have faired any worse. There is a reasonable statistical argument to
demonstrate that 400 trials are a good limit.
Buffer Screening
Anil Mistry
To find the best buffer system which will keep your protein happy, stable, and
soluble for concentration prior to crystallisation, setup your protein (at 1-2 mg/
ml) in hanging drops over 1 ml well solutions containing a series of buffers at var-
ious pHs, with various additives/stabilizers, etc. (but no precipitant!). Checking
for drops which are clear will give an idea of which solutions keep the protein
happy. In addition, as the system attains equilibrium some in situ concentration
of protein will be induced due to the bulk difference between the drop and
well volumes, hence an idea of how the protein behaves upon concentration
in this solution will also be observed. Modifying a clear or slightly clear drop by
adding a higher concentration of buffer to the well may even produce crystals.
However, the main piece of information this method can produce is an idea of
which buffer system and additives to put your protein into prior to concentration
a crystallisation screening.
Soft Crystals
Allan DArcy
If you have crystals that are very sensitive to being touched (they break) or stick
to the glass or dish, use a pointed strip of parafilm to move them. Otherwise,
grow the crystals on parafilm and punch wells around the crystals to move it.
Crystal Annealing
Clare Stevenson
As I said in my talk, give it a go. You might be surprised!!
No More 4C
Marie Anderson
Prepare trays for crystallisation, leave at 4C, and fill polystyrene box or flat
container with ice. Imbed a metal plate in the ice and set out cover slips. When
youre ready to set out crystallisations place the trays in the bed of the ice and
prepare drops, when finished transfer to 4C. Simple but it works.
Cross Seeding to Generate Crystals of a Related Protein or Protein/
Inhibitor Complex
Margaret OGara
If your protein or protein complex fails to crystallize try seeding from crystals of the
same protein (if its a protein you want to crystallize) or a related protein for apo
protein crystals. Serial micro seeding works best, make sure you look at the drops
after seeding to identify any visible crystal seeds in there (i.e. not new crystals).
Advice for the Follically - Challenged Crystallographer
Jonathon Hadden
It has been long accepted that crystallizers with an abundance of facial hair
have highly successful careers (Leeds University personal observation). This was
thought to stem from the fact that matter found its way from the hair into the
trials and acted as a nucleation centre-BUT SERIOUSLY; Addition of small grains
of sand to a crystallization drop that is close to producing crystals can aid in
nucleation and crystal growth.
Raise the DMSO?
Melissa Harris
If higher DMSO does not damage your protein, try higher concentrations of
DMSO (10-20%) for crystallization. It can help when dealing with insoluble com-
pounds and is an excellent cryoprotectant. Freeze directly from drop!
TCEP as a Reducing Agent
Barbara Brandhuber
Use TCEP instead of DTT as a reducing agent in your protein solution. It isnt
oxidized as quickly as DTT. Be sure to watch the pH of your solutions because
it is very acidic.
Temperature Variation
Irene Weber
To grow crystals at different temperatures around room temperature search the
lab for spots that are consistently at higher or lower temperatures. A difference of
several degrees can be found. Temperature shifts can be easily made by moving
crystals to a different place. (Check office shelves too!) [Discovered by Charles
Reed in my lab]
Drop Drying Technique
Anil Mistry & Ron Rubin
When setting up drops with a protein which has low solubility a low starting con-
centration has to be used. Setup larger drops (5-10 l) and leave them to stand
dry for 3-5 minutes at room temperature /4C prior to inverting over a well of
a Linbro

plate, this should allow some pre-concentration.

P A G E 3 - 7
Solutions for Crystal Growth
Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com 216 Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com
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Using DLS to Test for Irreversible Aggregation
Anil Mistry
Generally, people concentrate a protein to 10 mg/ml or higher, then dilute 2-fold
when setting up their hanging drops by doing a 1:1 minx. What if in concentrat-
ing to 10 mg/ml aggregation has been induced which is irreversible, such when
pipetting your 1:1 mix hanging drop, it already contains aggregates, a bad starting
point.
(For monodisperse protein samples)
Using DLS test the limit of concentrated protein, i.e., the maximum concentra-
tion that can be achieved before a polydisperse signal is obtained. Then test
samples, up to this limit, by concentrating up to this limit and test for irreversible
aggregation by diluting a concentrated sample to a number of levels and test for
monodispersity. With the sensitivity of current DLS equipment even samples at
10 mg/ml should be measurable. In this way and within the limits of your DLS
machine it should be possible to find out whether you will have aggregates when
you dilute your concentrated protein 2-fold when setting up a hanging drop.
Pickled Crystals Unusual Additives! (A True Story)
Michael Hickey
Pickle juice was added as an additive to a mutant form of a crystallized native
protein. The mutant could not be crystallized in near similar conditions of the
native. By chance, the components of pickle juice were read and found to con-
tain compounds used in crystallization (i.e. glycerol, PEG 400, citric acid, acetic
acid, alum, and a few vitamins). This juice (Sweet & Snappy Vlassic brand) was
filtered (0.45 l and adjusted to neutrality. It was then added to various PEGs
(that crystallized the native form) and set up with the mutant. Crystals formed
after 1 week! Trying to add back single components of the pickle juice to deter-
mine which component was responsible gave no crystals, the pickle juice (~1%)
was necessary. Hint/ Tip: Commercially available food/ detergent solutions ought
not be discounted as additives for crystallization!
Improve Your Crystals in Size, Shape, and Quantity
Nham Nguyen
After you have crystals, open the coverslip, remove the mother liquid in the
droplet, dissolve the crystal with 3 l of H20 or buffer and add 3 l of well solu-
tion. Close the coverslip. The crystal will appear again. My crystals diffract ~ 4 A,
sometimes I get twins that diffract ~2.5 A.
No Hits! What Next?
Anil Mistry, Pfizer
No hits from a screen, what next? You can always set up another screen! Or,
you can make a list of all drops that have precipitate and use the precipitate as a
potential seed stock. Streak seed from the precipitate into new drops that have
been set up at 50% and 75% of the original screening solution. There may be
nucleation sites or microscopic seeds in the precipitates that may grow at lower
precipitate saturation. Better still, streak seed from the precipitate to a clear zone
into the same drop to recycle the drop.
Micro-Seeding With A Cryoloop
Anna Aagaard, AstraZeneca
For reproducible micro-seeding by hand use a cryoloop to fish out your seed
from the seed stock and transfer them to the drop. Use a 0.3-0.4 mm cryoloop.
Reduce To Enlarge
Zhanna Druzina, SGX Pharma
For bigger crystals try to add 0.5-1 microliter of 14 M beta-mercaptoethanol to
the reservoir after the protein drop was set up.
Complex Screening
Annie Hassell, GlaxoSmithKline
Problem: Can grow crystal but no protein-ligand crystals. Tip: Take the condi-
tions from the apo crystals and develop a focused optimization screen (24 well
maximum). Screen complexes using cross seeding and the focused screen and
three drop ratios (1:1, 2:1 and 2:3).
Change the Method
Annie Hassell, GlaxoSmithKline
Problem: Poor crystal quality. Tip: Change tray type or crystallization method.
For example, initial screens done in sitting drop tray and crystal quality improved
in 96 well hanging drop tray.
Watch Out For Ruts
Brandon Collins, Boehringer-Ingelheim
Every project, every protein, every construct is unique. Be careful of knowing too
much. Just because things did or did not work in the past does not mean things
will work that way for the next project.
Heavy Atoms To The Rescue
Problem: Poor diffraction. Tip: Heavy atom soaks to stabilize floppy regions of
the protein.
Dont But All Your Eggs In One Basket
Joseph Luft, Hauptman-Woodward Institute
Don't focus all of your optimization efforts on a single crystallization condition.
If you have several different crystallization conditions identified for a sample
go after them. Crystals of the same protein produced from different chemical
conditions and/or temperatures will have unique physical properties. These
properties will determine how easy the crystal can be looped (physical stability),
cryoprotected and ultimately how well the crystal diffracts X-rays. Avoid single
points of failure, go after several hits.
P A G E 4 - 7
1991-2008 Hampton Research Corp. All rights reserved. Printed in the United States of America.
This guide or parts thereof may not be reproduced in any form without the written permission of the publishers.
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Insoluble Ligands
Doug Marcotte, Biogen IDEC
If ligands can't be soaked into crystal or co-crystallization is ligand specific try
seeding into drops that contain ligand of interest. When soaking crystals with
insoluble ligands try adding the cryoprotectant to the soaking solution. This can
help solubilize the ligand and also cryoprotect (so less handling). Doesn't work
so well when salt is the cryoprotectant, but may well when it's glycerol, DMSO
or ethylene glycol.
Getting Away From Clear Drops
Annie Hassell, GlaxoSmithKline
Problem: Drops are all/mostly clear. Tip: Remove stabilizing agents (salt, glycerol,
etc) from the protein buffer. Then do crystallization screens.
Avoiding Excessive Precipitation
Vaheh Oganesyan, MedImmune
At suboptimal protein concentrations the interface between protein solution
and crystallization screen solution may exhibit excessive precipitation. To avoid
this, before adding screen, solution add 1 microliter of water. Downside of this is
equilibration will take slightly longer. Upside of this is decreased osmotic shock
for protein and less precipitation.
Heavy Precipitate Control
Janet Newman, CSIRO
Put one conditions of 40% TCA into your standard screen. This should precipi-
tate out all of your protein, so that you have an idea of what heavy precipitate
should look like.
Look At The Big Picture
Edward Snell, Hauptman-Woodward Medical Research Institute
Don't consider a crystallization result in isolation. Look for neighbors in chemi-
cal space and use those results to provide chemical directions for optimization.
If a cryocooled crystal does not diffract well. You cannot tell if it is the crystal,
cryoprotectant or cooling that is causing the problem. Look at room temperature
data before moving on.
Poor Nulceation
Mei Xu, Novartis
Consider the case of poor nucleation and seeding did not work. Tip: Mix protein
and well solution then use pipette tip to cross the drop into branched shape.
Crystal may grow in the branches of the drop.
Seeding Suggestion
Paris Ward, GlaxoSmithKline
To increase you choices of producing more optimal crystal condition or condi-
tions using seeds, try the following. Program a small volume liquid handler to
dispense your protein and seed solution directly into 96 well commercial screens
and/or an additive screen.
Delete To Succeed
Heidi Schubert, University of Utah
Recent success with loop deletions. Sequence alignments reveal either charged
loops and/or loop insertions relative to homologues. I have removed 3 to 34
amino acids and retained high expression soluble protein and novel crystals.
Seeking Stability
Annie Hassell, GlaxoSmithKline
Problem: Unstable protein. Tip: Add 1-5% low molecular weight (200 to 1,000)
PEG directly to the protein and then screen.
Doh!
Jim Pflugrath, Rigaku
Start with big crystals! Add at least 5% glycerol to everything.
Stabilize For Cryo
Laura Pelletier, SGX Pharmaceuticals
Stabilize crystals in cryo by adding protein buffer components into the cryo. For
example, most commonly I add 100 to 150 mM NaCl plus reservoir components
plus cryoprotectant(s).
Ratios
Ayse Sinem Ozyurt, SGX Pharmaceuticals
Play with protein-mother liquor ratio, especially with low solubility proteins. Try
different concentration of protein coupled with streak seeding.
Eliminate One Offs
Nancy Bump, Millennium Pharmaceuticals
After looking at the results of initial screening or of additive screen, pick several
of the best "hits" and screen in 96 well format, 6 or 8 identical drops of each
favorite before scaling up. Helps to eliminate "one offs" and save time.
Complex It!
Paul Reichert, Schering Plough Research Institute
Apo protein is monidisperse but won't crystallize. Complex it! Complex it!
Complex it!
Matrix Seeding Tweak
Armando Villasenor, Roche Palo Alto
If your crystal seeds withstand large serial dilutions, try matrix seeding via the
reservoir by doing the following. 1) Create crystal seeds as described by Allan
D'Arcy. 2) Dispense seed into reservoirs containing reservoir. 3) Aispirate/
dispense to mix seed in reservoir. 4) Dispense mother liquor droplet containing
seed onto protein drop.
P A G E 5 - 7
Solutions for Crystal Growth
Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com 218 Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com
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A Little Salt Please
Neil Grodsky, Pfizer Global Research & Development, La Jolla Labs
If you do not see any crystal growth in several days after set-up (more than 1 to
2 weeks) and the drop are not all clear, add salt (such as ammonium sulfate) to
0.5 M to the drops. Even though ammonium sulfate salt crystals might form, you
might actually get protein crystals. This worked for me recently.
Urea To Solubilize
Gloria Borgstahl, Eppley Institute UNMC
Non denaturing (less than 3 M) concentrations of urea can be helpful to solubi-
lize your protein.
Multivalents
Simon Low, Pfizer, Inc.
Try stoichiometric levels of multivalents, cations as additives. They may be neces-
sary for crystallization but sometimes the levels found in commercial screens are
too high and toxic.
Cooking for Crystals
Liping Wang, GlaxoSmithKline
Heat treatment of protein complex to obtain diffraction quality crystals. Original
complex has no initial crystal hits. Heat treat protein complex (25 to 80 degrees
Celsius) for various times (5 to 30 minutes). Centrifuge to get rid of aggregated
protein and screen again.
Toothpick Tip
Paul Reinfelds, Illinois Institute of Technology
When using pre greased trays, take a toothpick and remove a bit of grease from
each well. Now as you push down on your cover slip, you turn it a few degrees.
This will allow the air to escape and the turn will form an airtight seal over each
well.
Thermofluor To The Rescue
Jackie Day, Pfizer St Louis
Thermostability. Monitor the effect of additives, buffers, ligands, etc. on melt
temp of your protein. We have seen in multiple cases that the most thermostable
construct, buffers, additive yields the best or only crystals. How? We use Bio-Rad's
iQ5 iCycler (a PCR instrument) as it has 5 sets of filters for excitational emission
and hydrophobic dyes that fluoresce upon binding (protein unfolding).
Getting Out Of A Sticky Situation
Michael Wiener, University of Virgina
Problem: Crystals adhering to plastic of sitting drop plate, and mechanical
dislodging (by cryoloop, tool, etc) does not work. Solution: Stan a fine gauge
syringe needle into the plastic, near but not into the crystal. This often distorts/
disrupts the plastic near the crystal and breaks the seal.
Liquid Bridge
Margarete Neu, GSK Stevenage, UK
Getting bigger crystal by low tech / low cost counter diffusion. If you are faced
with either no nucleation or showers of crystals and the usual tricks including
seeding do not work, try this: On a cover slide, set the protein and precipitant
drop (example 1 microliter plus 1 microliter) separate, but very close to each
other. Then, with a whisker or pipette tip streak through the drops to form a
connecting bridge between the protein and precipitant solution. Invert cover
slide and place over well. Crystals will form along the gradient and "self screen"
for best conditions.
Low Concentration Complexing
Elizabeth Fry, Abbott Laboratories
When working with compounds for co-crystallization, if the compounds are
highly insoluble in protein buffer (50 micromolar or less) we often employ low
concentration complexing. We dilute the protein and then add in diluted com-
pound, so tha the compound is added close, or at least closer to a concentration
where it is soluble and the content of DMSO in the protein sample remains
less than 2%. The protein-compound complex is then concentrated for crystal-
lization trials. This has helped us with several projects with highly insoluble
compounds.
Ionic Liquids
Christopher Bonagura, Exelixis, Inc.
In experiments using model proteins we found that Ionic Liquids (IL's) specifi-
cally 1-Butyl-2-methyl imidazolium chloride gave increased numbers of crystal-
lization outcomes compared to the IL controls. A large number of the crystals
obtained had precipitated outcomes in the IL controls. In many other cases the
IL and crystal had an improved morphology (needles to plates, plates to 3D crys-
tals) over the IL controls. Tip: Using an IL such as 1-Butyl-2-methyl imidazolium
chloride as an additive to improve chances of getting a crystal from conditions
which otherwise would give precipitate. Marc Pusey, MI Research, Inc. Our favor-
ite cryoprotectant. 1x UCP (Ultimate Cryo Protectant) 8% Glycerol, 8% Ethylene
glycol, 9% Sucrose, 2% Glucose. We make a 2x solution. Generally add this 1:1
with reservoir. The ratio can be modified, for example 1.2 microliter 2x UCP :
0.8 microliter reservoir, or 1.4 microliter 2x UCP : 0.6 microliter reservoir, or 0.6
microliter 2x UCP : 1.4 microliter reservoir, etc. I believe this has been success-
ful in cryoprotecting some 70% of all of our systems, resulting in more than 50
solutions of these targets regardless of previous cryogenic treatments. Author in
unknown to me, but the credit is published in a singled Hencrickson paper. I
was tipped off 6 years ago.
P A G E 6 - 7
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Protect The Bridge
Shirley Robert, York Structural Biology Laboratory
If you can't get your Se-met protein to crystallize try leaving out the DTT/TCEP
during purification and crystallization. Sometimes there are disulfide bridges
near crystallization contacts that need preserving for crystallization to take place.
Collected the Se edge is still possible.
DMSO For Cryo
Rich Romero, SGX Pharmaceuticals
Try 20 to 30% DMSO as cryo. This has worked well in a number of cases for me
and I've added this to a very short list of cryos that I personally use. Note: If your
mother solution contains a high salt concentration the DMSO will cause it to
precipitate out of solution. So beware!
No Fog
Beat Blattmann, University of Zurich
4 degrees Celsius crystallization plates prepared at room temperature always
have a condensation problem on the plate seal. To avoid condensation cover
the finished plates with two lids and plate it in the cold room for 20 minutes or
on top of a cold metal block, The will reduce the temperature of the reservoir
solution while the 2 lids delay the temperature change from the top long enough
to avoid condensation.
Anti Slip Tip
Barbra Pagarigan, Celgene
Ever been manually sealing a plate only have it slip out from under you? The
result is usually death for hanging drop plates, and with sitting drop plates your
best bet is hoping the drop is not splashed onto the seal above. To ensure the
plate stays put when applying pressure, we use a "grip pad" in our lab. Simply
place the grip pad onto the bench, set plate on top and seal as usual. The grip
pad prevents the plate from slipping out from under the compression tool, usu-
ally a brayer, used for ensuring the please seal is applied correctly. The grip pad
can be cut from the material commercially available for lining tool shop drawers.
In addition, we created a fixed plate holder that encloses the entire plate to guar-
antee the brayer does not slip off the plate when sealing, a common occurrence
when manually sealing many plates. Our plate holder is custom cut from a hard
rubber to fit both thee 24 well and 96 well plates. The plates sit slightly above the
platform to ensure both ends are sealed and for easy removal. The sides of the
platform are rounded to ensure the brayer has a smooth path of travel.
P A G E 7 - 7
Solutions for Crystal Growth
1991-2009 Hampton Research Corp. All rights reserved. Printed in the United States of America.
This guide or parts thereof may not be reproduced in any form without the written permission of the publishers.
cryst alli zati on ti ps from a to z
Crystals of apotransferrin (bovine milk) grown in fumaric acid, pimelic acid, sebacic acid, hexadecanedioic acid, maleic acid,
glutaric acid, suberic acid, dodecanedioic acid, oxamic acid, 15% w/v polyethylene glycol 3,350.
Peter Nguyen, Hampton Research, USA.
221
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P A G E 1 - 1 4
A is for Aggregation
Aggregation can be a deterrent to the crystallization of biological macromol-
ecules including proteins, peptides, and nucleic acids. The presence of sample
aggregation can be detected by either dynamic light scattering or native gel
electrophoresis. Aggregation might be caused by hydrophobic patches on the
surface of the sample, differently charged isoforms, differently phosphorylated
isoforms, mixtures of methylated and non-methylated samples, glycosylation, as
well as electrostatic interactions. Aggregation can be due to autologous aggrega-
tion where the protein is aggregating with itself or heterologous contamination
where the sample is aggregating with other proteins. In the case of heterologous
contamination, further purification of the sample should be seriously consid-
ered. In the case of autologous aggregation that precludes crystallization, one
might consider:
Using molecular biology to manipulate intra and inter molecule interactions by
modifying the sample sequence (alter, add, or delete residues).
Using chemical additives, like those listed below, to manipulate
sample-sample and sample solvent interactions:
Detergents
Chaotropes (urea, guanidine hydrochloride, hydrochloric acid, etc.)
Electrostatic agents
Alcohols (isopropanol, methanol, ethanol, etc.)
Salts (sodium chloride, potassium chloride, sodium fluoride, etc.)
Polyols (glycerol, PEG 400, etc.)
Ligands, inhibitors, co-factors, and metals
Use temperature to prevent aggregation (0C and 60C)
Consider a fusion protein
Remove C-terminus or N-terminus
Truncate domains
Remove His-tag
In some cases aggregates can be removed by centrifugation or filtration.
In some cases the aggregates can be removed by mixing the sample with the
crystallization reagent, allowing the sample to incubate for 15 minutes, centrifug-
ing the sample/reagent mixture, removing the precipitate, and setting the drop
with the supernatant.
B is for Birefringence
The technical mumbo jumbo first. The physical properties of isotropic materials,
such as glasses, liquids, and amorphous materials, do not depend on direction.
However, most properties of a wide variety of crystals (including liquid crystals)
do show such variation. This anisotropy of physical properties originates in the
anisotropic build-up of the materials (crystal structure). Anisotropy in the optical
properties of uniaxial crystals is referred to as either birefringence or dichroism,
depending on whether the index of refraction or the absorption coefficient is
concerned. Birefringence means that there are two distinct speeds with which
light can propagate, depending on the direction of propagation. When a light
ray splits into two beams as it passes through a material, the effect is called
birefringence (or double refraction) and the material is birefringent. If you look
at something through a birefringent material, youll see double. The word bire-
fringence comes from the Latin bi- (twice) plus refringere (to break up). Thus,
the light rays are broken in two by a birefringent material. One well-known
example of a birefringent medium is crystalline calcite (calcium carbonate). If
you look at the world through a clear crystal of calcite (calcium carbonate), you
will see double. Place such a crystal on a drawing, and youll see two overlapping
copies of the drawing. The molecular structure of calcite causes double refrac-
tion, in which each light ray is split into two rays that emerge from the crystal at
slightly different angles. Calcite shows this more clearly than most crystals, but
quartz and many other crystalline minerals also split light ray.
Now, the practical interpretation for crystal growers. You might hear the word
birefringence used quite often by crystal growers when viewing crystals under
a microscope. Here, crystal growers are stretching the definition of the term
birefringence to describe the colorful display produced by biological macromo-
lecular crystals when polarized light is passed through the crystal.
A light microscope with polarizing optics is required to observe birefringence.
The following path is a typical setup. Light passes from the light source through
the first polarizing lens, then the specimen (crystal) then the second polarizing
optic, the magnifying optics and then into your eye. On many typical polarization
setups, the second polarizing filter can be rotated while the specimen is station-
ary. Rotating the polarizing optic without something to rotate the plane of polar-
ized light in the path (i.e. a crystal) will result in one seeing light, dark, light, dark,
as the filter is rotated. But if a crystal with birefringent properties (i.e. a biological
macromolecular crystal) is positioned in between the two polarizing filters, one
will observe changing colors as the polarizing filter is rotated. Specifically, when
the polarizing filters are aligned such that the field is dark, a birefringent object
(crystal) will glow with color.
Birefringence is one way we can differentiate amorphous precipitate from micro-
crystals in a drop when viewed under a microscope. Precipitate does not have
birefringent properties while most biological macromolecular crystals do.
One drawback with using birefringence in todays crystal growth world is that
most of the crystallization devices utilized are made from plastic such as polystyrene
and polypropylene. These plastics are optically active and can be birefringent. In
fact, often times the colors we see displayed in crystals are contributions from
the plastic birefringence. However, it is still possible to observe microcrystalline
birefringence in the plastic trays, but there is usually a contributory effect from
the plates used to grow the crystals. One way to avoid this is to grow crystals in
a glass device or at least observe the crystals in a path that is free of birefringent
plastics.
Birefringent precipitates will glow, sparkle, or glisten.
To test for birefringence, position the polarizers so the field of view is dark
WITHOUT they crystallization plate or setup. Place the tray into position on the
microscope. If a crystal is birefringent, some of the light passing through the
crystal will be rotated and pass through the second (analyzing) polarizing filter.
The intensity of the transmitted light will increase and decrease as the crystal or
the polarizer is rotated. Remember, birefringence is not ALWAYS clearly visible
when plastic is in the light path (i.e. when you use plastic slides or crystallization
plates). However, a birefringent crystal viewed in a plastic tray or plastic cover
slide will have a different color than the background (i.e. plastic plate) or fore-
ground (plastic slide). Finally, birefringence is a property of crystals, both
crystalli zati on
ti ps from a to z
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P A G E 2 - 1 4
biological (proteins, peptides, and nucleic acids) and inorganic crystals (salts).
Birefringence is MORE pronounced in inorganic crystals (salt).
A quick comment on what to do with birefringent precipitates. Streak seeding is
a common and often successful method of taking advantage of microcrystalline
precipitate to grow large single crystals. But that starts with an S so we cannot
talk about that this time!
C is for Condensation
Condensation on cover slides or tape may be a problem when incubating plates
at cool or warm temperatures.
To minimize condensation try the following: Place a dummy plate above and
below the plates being used for crystallization. For example, on a stack of 5 or 10
plates, place a dummy plate on the bottom of the stack as well as the top of the
stack. The reservoirs (wells) of the dummy plates should contain water as the
reservoir solution. The reservoirs should be sealed either with plain glass cover
slides and vacuum grease or clear sealing tape to prevent the reservoir solution
from evaporating.
Pipet the reservoirs and drops at the temperature where the plates will be
incubated. This means enjoying a cold room during setups. One can cheat and
avoid a cold room by incubating reagents, pipet tips, plates, and slides at the cold
temperature. When ready to set the experiment, set the plates in an ice bath and
quickly set the plate while the plate (as well as reagents and protein) are on ice.
Conceivably one could also do this for warm temperature incubations where an
elevated temperature incubator is available. The idea is to avoid a RAPID change
in temperature with the sealed crystallization experiment (plate). By minimizing
the temperature difference between the setup and incubation temperatures,
condensation can be minimized.
Try sitting drop vapor diffusion. Condensation is especially bad for hanging
drop vapor diffusion experiments since water condensing on the slide can run
or merge into the crystallization drop. To avoid such a merger, try sitting drop
vapor diffusion. Condensation on the tape or cover glass above a sitting drop
experiment may still make viewing the drop difficult, but at least the drop will
not become diluted with condensation unless the condensation is so bad that it
rains in your experiment!
Finally, avoid vapor diffusion altogether and go microbatch.
D is for Drafts
Temperature is often a significant and overlooked crystallization variable.
Temperature can sometimes be an unwanted crystallization variable in labs
subject to temperature changes. Temperature fluctuations are especially trouble-
some because unless they are intentional and accurately produced and recorded,
the temperature changes are difficult or impossible to reproduce. Keep the fol-
lowing in mind when selecting a location to store crystallization experiments:
How does the temperature of the room fluctuate over the course of a day, over
the weekend, during the month, over the year?
Place a thermometer in a container full of water (allowed to equilibrate to the
location temperature) and observe the temperature fluctuation over time to
determine the temperature stability of the location.
Is the area subject to drafts such as opened and closed doors, under a heating or
air conditioning duct, or near a heat producing device?
Avoid storing experiments near windows or walls that are in contact with the
outside of the building.
Is the heating or air conditioning turned off in the evening or on weekends in
your facility? If yes, substantial temperature changes can occur, especially during
very cold or hot days/nights.
In an effort to maintain a more constant room temperature, store the plates
in a quality, commercial incubator set at the desired room temperature. Since
most of us cannot swallow spending money for a room temperature incubator,
consider storing the plates in an unplugged or non-working refrigerator, styro-
foam box, ice box, or cooler.
Use a quality incubator to store crystallization experiments.
E is for Ethylene Glycol
Ethylene glycol. Also known as 1,2-Ethanediol as well as glycol. Molecular mass of
62.07. HOCH
2
CH
2
OH. A nice little polyol that can be useful as a cryoprotectant.
Typically used as a cryoprotectant in the concentration range of 10 to 30% v/v.
Also useful as a non volatile organic additive when used in the concentration
range of 1 to 5% v/v and in some cases, much higher (15 to 25% v/v). As an
additive, ethylene glycol has been reported to sometimes increase the size
and quality of the crystal. Finally, it is useful as a crystallization reagent in the
concentration range of 25 to 30% v/v. It is even thought that ethylene glycol can
have a structure-stabilizing effect and may be useful in the crystallization of
flexible proteins.
F is for Fab Fragments
Fab and Fv fragments can be used as co-crystallization agents.
Fab fragments have decent solubility properties and bind specifically to antigens
with reasonable equilibrium constants (105 to 108 M-1). Fab fragments can, in some
instances, effectively transform aggregated protein into soluble, monodisperse pro-
tein, suitable for crystallization trials. Since antibodies are frequently available from
related biochemical studies, they can often be applied more readily than molecular
biology (point mutants, truncations, molecular engineering). However, it seems
MOBO gets faster and easier with each passing year. It is often useful to consider
screening several different Fab fragments that recognize different epitopes for
crystallization trials. Sometimes Fab fragments can immobilize a region of a protein
sample, reducing sample flexibility, enhancing conformational homogeneity of the
sample which in turn can enhance chances for crystallization.
Using an Fv fragment in lieu of a Fab fragment might have some advantage since
there is no flexible elbow to inhibit crystallization.
When preparing antigen-antibody complexes for crystallization, one should
carefully select the antibody, prepare a homogeneous Fab species, and prepare
the Fab-sample complex with proper stoichiometry. Ideally the Fab should not
interfere with the native sample conformation or activity.
The presence of a Fab fragment in the crystal can assist in the crystallographic
structure determination by allowing one to utilize molecular replacement.
Fab and Fv fragments should be considered useful tools in the crystallization
cryst alli zat i on ti ps from a to z
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toolbox for manipulating sample solubility, conformational flexibility, and crystal
lattice contacts, as well as being a possible tool for molecular replacement.
Reference:
The use of antibody fragments for crystallization and structure determinations. LC Kovari, C. Momany and MG Rossman.
Structure 15 December 1995, 3: 1291-1293.
G is for Gap
When greasing crystallization plates, it is often useful to leave a small gap in the
bead of grease applied to the plate. For example, begin greasing at 12:00 and
apply the bead clockwise until 10:00. A gap with no grease is left between 10:00
and 12:00. Place the cover slide onto the bead of grease. Depress the slide onto
the grease. As the slide is pressed onto the bead of grease, pressure is allowed to
escape through the gap. Before the slide bottoms out on the plate, twist the slide
approximately 15 minutes (90) to smear the grease across the gap and properly
seal the cover slide to the plate.
H is for 1,6 Hexanediol
1,6 Hexanediol (C
6
H
14
O
2
M
r
118.18) is a non-volatile alcohol which most likely
precipitates biological macromolecules by lowering the chemical activity of water.
Non-volatile organics can be effective crystallization reagents for proteins and are
particularly appropriate for nucleic acids. This non-volatile alcohol steals water
molecules from the biological macromolecule through hydrogen bonding and
reduces the electrostatic screening effectiveness of the water. Typical stock solu-
tions of 6.0 M (about 71% w/v) are typical for this crystallization reagent. Typical
reservoir concentrations for this reagent are 1 to 4 M across a broad pH range.
1,6 Hexanediol can be utilized in the presence of numerous anions and cations,
as well as a broad range of salts. It can be a substitute crystallization reagent for
MPD, PEG 400, and PEG MME 550. It can also be utilized as an additive in the
concentration range of 0.1 to 0.2 M. 1,6 Hexanediol is a waxy solid or flake. When
formulated with water, the reaction is endothermic and one might need to warm
the solution to effect complete dissolution at high levels of supersaturation.
1,6 Hexanediol is utilized as a precipitant in Crystal Screen 2 and is available as a
preformulated, sterile filtered reagent from Hampton Research (Catalog Number
HR2-625; 6.0 M 1,6 Hexanediol, 200 ml).
I is for Interface Diffusion
Crystallization by interface diffusion was popularized by Ray Salemme (F.R.
Salemme 1972, Arch. Biochem. Biophys. 151, 533). The method is also called
free interface diffusion and liquid-liquid diffusion. The methodology is straight-
forward. Interface diffusion is classically performed in a cylinder with a small
diameter, such as a capillary. Using a capillary with both ends open, the sample
is pipetted into the capillary. Next, the crystallization reagent is added to the
capillary. For true interface diffusion to occur, the sample and the reagent should
touch one another inside the capillary. This can be accomplished by making sure
the sample is positioned at the inside end of the capillary before adding the crys-
tallization reagent. When adding the crystallization reagent, use a pipet tip with
a small diameter opening such as a gel loading tip. Carefully and slowly pipet
the reagent from the pipet tip and allow a bead of the reagent to form at the tip.
Hold the capillary parallel to the ground and gently touch the tip of the capillary
to the reagent bead. With a combination of gently pipetting, capillary action and
proper capillary tilt, fill the capillary with the appropriate amount of crystalliza-
tion reagent without air gaps. Done correctly, the inside of the capillary should
contain a single cylinder of liquid. Filling the capillary should be done gently and
slowly to minimize mixing the sample and the reagent. Actually, one should add
the more dense solution into the capillary first, followed by the less dense solu-
tion to minimize mixing. If done properly, one can see an interface between the
sample and the reagent. It might be best to start with equal amounts of sample
and reagent. Later, one can vary the ratio of sample to reagent for optimization.
Once filled with reagent and sample, the capillary can be sealed with a non-dry-
ing clay or wax (sealant). Be careful not to add too much sealant at a time to
the end of the capillary since the sealant will displace the liquid in the capillary
and too much sealant will result in liquid being lost from the capillary. Storage
of the capillary can be done in several ways. Some like to simply stick the capil-
laries vertically into clay or wax. But this means one must remove and contort
the capillary for viewing under a microscope. Instead, one can use a petri dish
(square ones work really well) and line the petri dish with two thin lines of wax
or clay about the size of a toothpick. The lines of wax or clay should be parallel to
one another and positioned apart from one another just under the length of the
capillary. The capillaries can be set upon the lines of clay in neat rows. A dozen
or so capillaries can be stored in a small square petri dish. Storage and viewing is
quick and simple. The petri dish also facilitates easy labeling.
A nice feature of interface diffusion crystallization is the potential of a concentra-
tion gradient of sample and reagent along the length of the capillary. Although
pipetting, gravity, and movement make this interface and gradient less than per-
fect, gradients do form and one can often visualize the concentration gradient as
crystals of different size and number form at different points along the gradient
(length of capillary), presumably due to the change in relative supersaturation
along the length of the capillary. For example, numerous, small crystals might
form at high levels of supersaturation, fewer and larger crystals might form at
ideal levels of supersaturation, and no crystals may form at very low levels of
supersaturation.
Permutations of interface diffusion include the following. Doing it in micrograv-
ity. Freezing one layer before adding another to ease pipetting (good for larger
volumes). Freezing the entire experiment then slowly melting the slug to intro-
duce temperature as a crystallization variable. Placing a gel plug between the
sample and the reagent. Using high throughput plates with tall, thin cylinder-like
wells for vertical interface diffusion.
J is for Jeffamine

Reagent
The Jeffamine polyoxyalkyleneamines contain primary amino groups attached
to the terminus of a polyether backbone. They are thus polyether amines.
The polyether backbone is based either on propylene oxide (PO), ethylene
oxide (EO), or mixed EO/PO. Jeffamines are synthesized as either monoamines
(M-series), diamines (D series), or triamines, and are made in a variety of molecu-
lar weights, ranging up to 5,000. The ED-series are aliphatic diamines structurally
derived from the propylene oxide capped polyethylene glycol.
The wide range of molecular weights, amine functionality, oxide type, and
distribution provides flexibility in synthetic design of compounds made from
Jeffamine Reagents. For the most part, Jeffamine

Reagent products
undergo typical amine reactions and are low viscosity liquids, exhibiting
low vapor pressure.
Solutions for Crystal Growth
Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com 224 Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com
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Jeffamine

Reagents originated chemically at the Texaco Chemical Company as

lubricants and fuel additives and are now most frequently used in manufacturing
adhesives, coatings, epoxies, and curing agents. Scary what we crystal growers
will use as crystallization reagents, is it not?
Jeffamine Reagents worked their way into the protein crystallization community
in the late 70s when Dulio Cascio (then in the lab of Alexander McPherson at the
University of California Riverside now University of California Los Angeles) evalu-
ated them as polymeric precipitants similar to the polyethylene glycols.
Alexander McPherson published the use of Jeffamine Reagents in a crystallization
publication Two approaches to the rapid screening of crystallization conditions
J Crystal Growth 1992, 122: 161-167.
The first protein structure solved using Jeffamine Reagents was that of Xylose
Isomerase in the publication by Lloyd et al, Crystallization and preliminary X-ray
diffraction studies of xylose isomerase from Thermoanaerobacterium thermo
sulfurigenes strain 4B J Mol Biol 1994, 240: 504-506.
Jeffamine

Reagents are discussed by Lesley Lloyd Haire and others in Terese
Bergfors wonderful book Protein Crystallization - Techniques, Strategies, and
Tips, A Laboratory Manual (IUL Biotechnology Series 1999 ISBN 98-075232).
The following Jeffamine Reagents have been tried in protein crystallization
experiments:
Jeffamine D-230 Reagents
Jeffamine D-400 Reagents
Jeffamine ED-600 Reagents
Jeffamine

ED-900 Reagents
Jeffamine ED-2001 Reagents
Jeffamine M-600 Reagents
Hampton Research has experienced the most success with Jeffamine ED-2001
Reagent and Jeffamine M-600 Reagent.
Jeffamine Reagents can be used like polyethylene glycols in the crystallization
of proteins, peptides, and nucleic acids. In fact, some crystal growers substitute
Jeffamine Reagents for PEGs during optimization to see if using them can
improve the crystals or offer new conditions for further optimization.
Jeffamine Reagents can be formulated with most of the salts, buffers, organic
solvents, and additives used in biological macromolecular crystallization.
There is very little literature describing the use of Jeffamine Reagents as crystal-
lization reagents and not a single report of them being used as a crystallization
reagent in the Biological Macromolecular Crystallization Database at the time of
this writing.
(http://wwwbmcd.nist.gov:8080/bmcd/bmcd.html)
Obtaining crystals in Jeffamine Reagents is a mixed bag. It is a good thing since there
are crystals. It is a bad thing since they are a pain to formulate.
The formulation of Jeffamine Reagents as crystallization reagents is tricky and
tedious. They are very alkaline chemicals and must be titrated to neutrality or
the desired experimental pH before use. Titration is typically performed using
hydrochloric acid. A significant amount of hydrochloric acid must be used to
titrate the Jeffamine Reagents to pH values between 2 and 11. pH changes are
slow until pH 9, and change very rapidly as one approaches neutrality. The
addition of hydrochloric acid results in a significant temperature increase in the
reagent. Accurate pH recordings require repeated pH, cool, pH, titrate; repeat
cycles. Many salts, including anions and cations, can be used in conjunction with
Jeffamine Reagents. Typical salt/Jeffamine Reagent ratios and concentrations are
similar to those used with related polyethylene glycols. Phase separation can be
a bigger problem with Jeffamine Reagent / salt mixtures than for PEGs, but can
be minimized by titrating the Jeffamine Reagent to neutrality or close to the final
working pH before adding the salt and/or buffer. Having as much water present
in the formulation also helps prevent phase separation.
Jeffamine M-600 Reagent is utilized as a crystallization reagent in Crystal Screen
2 (Catalog Number HR2-112).
Hampton Research offers the following preformulated, ready-to-use Jeffamine

Reagents:
Jeffamine ED-2001 Reagent 50% w/v solution pH 7.0, 200ml sterile filtered
(HR2-597)
Jeffamine M-600 Reagent 50% w/v solution pH 7.0, 200ml sterile filtered
(HR2-501)
Hampton Research also offers the following Jeffamine Reagent which has not
been titrated:
Jeffamine M-600 Reagent 100% solution pH >12, 200 ml (HR2-503)
Jeffamine is a registered trademark of the Huntsman Petrochemical
Corporation.
K is for Krypton
Named from the Greek kryptos or hidden, krypton is neither green, nor a solid
material that can defeat Superman. Rather it is another noble gas discovered in
1898 by Ramsay and Travers. It ranks sixth in abundance in the atmosphere.
Krypton gas is used in various kinds of lights, from small, bright, flashlight bulbs
to special strobe lights for airport runways. As with the other noble gases, kryp-
ton is isolated from the air by liquefaction.
Krypton and crystals. The use of xenon and krypton at high pressure is becom-
ing a popular method for the phasing of proteins. There are several homemade
as well as commercially available pressure cells for creating xenon and krypton
derivatives.
In essence, the crystal is exposed to the krypton or xenon gas at high pressures
for a short period of time in order to allow the krypton or xenon to bind to
the protein within the crystal. Following incubation and depressurization, the
crystal is flash cooled in liquid nitrogen and mounted onto a goniometer in a
cryostream for X-ray diffraction analysis. Freezing of the crystal is necessary to
prevent the diffusion, release and loss of gaseous krypton or xenon. As a side
note, methods of generating and analyzing xenon derivatives in glass or quartz
capillaries have been described (see references below).
Krypton and xenon binding sites are generally different from those for other
heavy atom sites so the screening of krypton and xenon can be used as a follow
up when the initial soaks in heavy atoms are not successful.
When the specialized hardware is available for screening krypton and xenon
cryst alli zat i on ti ps from a to z
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under high pressure, the method is a convenient and rapid way for screening
successful derivatives for phasing.
References:
High-pressure krypton gas and statistical heavy-atom refinement: a successful combination of tools for macromolecular structure
determination. Schiltz, M., Shepard, W., Fourme, R., Prange, T., de La Fortelle, E. and Bricogne, G. Acta Cryst. D53: 78-92, 1997
Exploring hydrophobic cavities in proteins using xenon and krypton noble gas. Prange,T., Schiltz, M., Pernot, L., Colloch,
N., Longhi, S., Bourguet, W. and Fourme, R. Protein Struct. Funct. and Genetics 30(1): 61-73, 1998.
Solubility of krypton and xenon in blood, protein solutions, and tissue homogenates. Yeh, S.Y., Peterson, R.E. J Appl
Physiol 5: 1041-1047, 1965.
Flash freezing isomorphous xenon or krypton derivatives of protein crystals. Sauer, O., Dutzler, R and Kratky, C. ECM 17,
Seventeenth European Crystallographic Meeting (Lisboa, Portugal 24/28 aug.1997). Book of Abstracts p 18 (ref. MS1.6-4)
Xenon and Krypton at LURE http://www.lure.u-psud.fr/sections/Xenon/XENON.HTM
Tutorial for Krypton-Elastase SIRAS refinement http://utica.med.jhmi.edu/sharp/tutorials/KrEl.html
L is for Lyophilization
Avoid lyophilization of your protein for storage and concentration if you plan to
use it for crystallization experiments. Lyophilization is a not-so-gentle procedure
and can prevent crystallization. Yes, we know that lyophilized lysozyme will
crystallize quite readily as well as other Sigma proteins but we have seen, as
well as others, many a protein that will not crystallize following lyophilization.
And we have yet to see a protein that would only crystallize after lyophilization.
So dont go there.
If you have a lyophilized protein, there is hope. First of all, one should solubilize
the protein in water or a stabilization buffer and dialyze the protein exhaustively
against water or the stabilization buffer. Dialysis is an important step which will
help to remove residual, non-volatile buffers and reagents, as well as low molecu-
lar contaminants.
L is for Lipidic Cubic Phases
Membrane proteins can be crystallized in a lipid phase where the crystallization
detergent diffuses into the lipid phase and crystal growth proceeds through
lateral diffusion of the protein molecules.
References:
Lipidic cubic phases: a novel concept for the crystallization of membrane proteins. Proc Natl Acad Sci 1996, 93: 14532-
14535
Related Reference:
X-ray crystallography of membrane proteins: Concepts and applications of lipidic mesophases to three-dimensional mem-
brane protein crystallization G PROTEIN-COUPLED RECEPTORS. 2000. p.365-388
L is for Light Scattering
Dynamic light scattering, also known as DLS, also known as photon correlation
spectroscopy, also known as quasi-elastic light scattering, can be utilized to
screen crystallization candidates for monodispersity.
DLS measures the translational diffusion coefficient of a macromolecular under-
going Brownian motion in a solution. What? In essence, DLS measures the inten-
sity of light scattered by molecules in solution. In turn, this measurement can tell
you the size distribution of the protein molecules in solution.
Crystal growers should use DLS to differentiate monodisperse solutions from
polydisperse solutions. Homogeneous, non-aggregated monodisperse proteins
have a high probability of producing crystals. Proteins with non-specific aggre-
gates and heterogeneous samples are less likely to crystallize. DLS allows one
to screen protein samples quickly, with small volumes of sample, at different
temperatures. The procedure is non-invasive so the protein can be recovered. If
DLS is not an option, one can obtain similar results using native polyacrylamide
gel electrophoresis or size exclusion chromatography. But DLS is faster and
more sensitive.
For a great story on DLS, as well as practical information on methods for
DLS, read Terese Bergors chapter on Dynamic Light Scattering in Protein
Crystallization - Techniques, Strategies, and Tips - A Laboratory Manual.
M is for Mass Spectrometry
Mass spectrometers (MS) use the difference in mass-to-charge ratio (m/e) of ion-
ized atoms or molecules to separate them from each other. MS is therefore useful
for quantitation of atoms or molecules and also for determining chemical and
structural information about molecules. Molecules have distinctive fragmentation
patterns that provide structural information to identify structural components.
MS can be a nice tool along with Dynamic Light Scattering (DLS), sodium dodc-
eyl sulfate polyacrylamide gel electrophoresis (SDS PAGE), iso-electric focusing
(IEF), and size exclusion chromatography (SEC) to evaluate the purity, homo-
geneity, and monodispersity of the sample prior to crystallization. Likewise, if
crystallization problems exist, MS can be a nice tool to identify possible sources
of the problem. MS can also be used to detect and possibly identify impurities
in crystallization reagents.
In the case of recombinant protein samples, an accurate measurement of the
molecular weight will inform one about the presence of post-translational modi-
fications. Dependent on the physical properties of the compound and the MS
technique used, the molecular weight may be determined within an accuracy of
one Dalton. The use of MS measurements will give sequence information where
classical methods fail or may lead to ambiguous results as in the case of N-termi-
nal blocked peptides, glycosylation or identification of the C-terminus. Next to
this, MS is the ideal analytical method to support sample clean-up procedures
and as a routine check for sample impurities of all kinds. As for sample require-
ments for MS, the samples should be free of non-volatile salts and the procedure
typically requires between 1 ug and several nmoles, depending on the kind of
analysis performed.
N is for Nucleus
For a crystal to grow, the system must be supersaturated, in non-equilibrium.
But first, a stable nucleus must form. A stable nucleus is an aggregate of the
macromolecule. A stable nucleus is an aggregate of such size, shape, and proper-
ties that it will enlist new molecules into its growing surface faster than others
are lost into solution.
N is for NIST/CARB BMCD
The NIST/CARB BMCD is a Biological Macromolecule Crystallization Database.
The internet address for this freely accessible database is:
http://wwwbmcd.nist.gov:8080/bmcd/bmcd.html
The BMCD contains crystal data and the crystallization conditions, which have
Solutions for Crystal Growth
Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com 226 Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com
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been compiled from literature. The current version of the BMCD contains 3547
crystal entries from 2526 biological macromolecules for which diffraction quality
crystals have been obtained. These include proteins, protein:protein complexes,
nucleic acids, nucleic acid: nucleic acid complexes, protein: nucleic acid com-
plexes, and viruses.
Use the BMCD to see if your sample has been crystallized or to see if a related
sample has been crystallized. Use this information to design crystallization
experiments for your sample.
N is for Nuclease Inhibitors
Nucleases can be a real pain during the crystallization of nucleic acids. Here
are some nasty things nuclease can do to your sample: Modification of sample
size, charge, or hydrophobicity, partial or total loss of activity, or utter desc-
truction of the sample. Traces of nuclease can be difficult to detect even when
overloading electrophoresis gels and can obviously cause sample damage
during purification, concentration, and storage of samples. What is one to do?
Well, one can include a nuclease inhibitor in the prep or sample to protect
the sample from ribonuclease and deoxyribonucleases. Inhibitors of nucle-
ases include RNasin (from Promega), ribonucleaoside-vanadyl complexes, and
DEPC. Inhibitors of deoxyribonucleases include DEPC and chelators such as
EDTA or EGTA.
O is for Osmium
Osmium is a good reactant for ribose moieties and the 3 terminus of RNA for
heavy atom derivatives.
O is for Organic Solvents
The most popular volatile organic solvents used in biological macromolecular
crystallization have been ethanol, acetone, isopropanol, tert-butanol, 1,3-pro-
panediol, acetonitrile, DMSO, methanol, and 1,3-butyrolactone. Organic solvents
can be utilized as a primary precipitant (buffered or unbuffered), as a secondary
precipitant in the presence of salt or polymer (primary precipitant), or as addi-
tives. The most popular nonvolatile organics have been MPD and 1,6-hexanediol.
Organic solvents act as precipitants by lowering the chemical activity of water.
This means they steal water molecules from biological macromolecules in solu-
tion, through a process of hydrogen bonding. This in turn reduces the dieletric
constant of the solution. Current popular thought is that organic solvents
should be used at low temperatures (4C or lower) and at the lowest possible
ionic strength, keeping in mind to include whatever is necessary to stabilize the
sample (buffer, divalent cations, etc.).
P is for Protease Inhibitors
Proteolytic modification of proteins can be a tool for making small active
fragments of proteins, which might have enhanced solubility characteristics
compared to the native protein, which in turn might make the protein more
amenable to crystallization.
However, proteases can also be trouble makers when they are uninvited guests
to a protein sample. When protease contamination can be a problem, one can
consider adding a protease inhibitor to prevent cleavage of the sample by these
nasty proteases. Protease contamination can occur when these nasty beasts are
carried over from isolation and purification of the sample from a natural source
or expressed proteins. Also, if your lab is situated in an area (on the same floor,
building, etc.) where microbial research, plant research, or other work involving
the possible generation of fungal and bacterial organisms is possible, then one
might consider adding protease inhibitors to the sample for protection. Why?
The presence of fungal or microbial contamination in your sample, reagents, or
crystallization related plates, capillaries, etc. can lead to growth of such organ-
isms with subsequent release of proteases from these organisms so they can use
your innocent little sample for food. Some crystal growers like to include sodium
azide or thymol in all their reagents and sample as a deterrent to microbial
growth, which prevents the possibility of microbial agents growing and secreting
proteases into the sample solution. But sodium azide and thymol can sometimes
bind the sample, are toxic, and in some cases do not live well with heavy atoms
so some feel it is not wise to simply include these agents as a multi-vitamin in
the crystallization cocktail. Rather, clean workspace, sterile filtered samples and
reagents, sterile pipet tips, and good technique go a long way to preventing
microbial contamination. Well, back to the P for Protease Inhibitors. Here are a
few protease inhibitors and their targets.
Protease: Metalloproteases
Inhibitor: Chelators like EDTA and EGTA, bestatin, amastatin, thiol
derivatives, hydroxamic acid, phosphoramidon.
Protease: Aspartic Acid Proteases
Inhibitor: Pepstatins and statin derived inhibitors.
Protease: Cysteine Proteases
Inhibitor: Thiol binding reagents, peptidyldiazomethanes, epoxysuccinyl
peptides, cystatins, peptidyl chloromethanes.
Protease: Most
Inhibitor: DEPC
Protease: Serine Proteases
Inhibitor: Trypsin inhibitors, leupeptin, boronic acids, cyclic peptides, DIFP,
PMSF, Pefabloc, aminobenzamidine, 3,4-dichloro isocoumarin,
chymostatin.
Q is for Quintillion
In the USA and France, quintillion is the number 1 followed by 18 zeroes. In
the UK and Germany, it is the number 1 followed by 30 zeroes. Use this term to
impress your lab mates, as in Hey, have you tried my new screen with a quintil-
lion different conditions? Just think of it, a screen with a quintillion conditions.
Gotta get a hit with that one. Problem is, most of us are lacking a quintillion mg
of protein...
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Q is for QELS (Quasi Elastic Light Scattering)
Another name for DLS (see D).
Q is for Quiet
Vibration has been reported to lead to excessive nucleation and crystals of
questionable quality. It is often suggested that crystallization experiments be
incubated in a location with minimal vibration. Avoid cabinets that are frequently
opened and closed, or countertops where equipment such as centrifuges or
vortexes live. Incubators can be another sore spot for vibration, especially poorly
insulated incubators with compressors that are not well isolated.
R is for Reducing Agents
Reducing agents are substances that cause other chemical species to be reduced
or gain electrons. In order for reducing agents to cause the gaining of electrons
on some other chemical species, they must undergo oxidation. Therefore reduc-
ing agents undergo oxidation when they do their job.
Dithiothreitol (DTT), beta-mercaptoethanol (beta-me), and Tris(2-Carboxyethyl)-
Phosphine Hydrochloride (TCEP HCl) are sulfhydryl protective reducing agents.
Reducing agents are typically used to prevent the oxidation of free sulfhydryl
residues (cysteines) in the protein. Such oxidation can lead to non-specific aggre-
gation of the sample, sample heterogeneity, inactivity, or denaturation of the
sample. In a typical crystallization experiment, reducing agents are used in the
concentration range of 1 to 10 mM in the crystallization drop.
Beta-me has one sulfhydryl group and is the weakest of the three reducing
agents discussed here, lasting perhaps two to three days. The supply of beta-me
should be replenished every two to three days in the crystallization experiment
to maintain the effectiveness of the reducing agent. Since beta-me is volatile, it
can be added to the reservoir of vapor diffusion experiments for diffusion into
the crystallization drop.
DTT has two sulfhydryl groups and lasts about three to seven days in a typical
crystallization experiment. DTT is not a strong volatile like beta-me (although it
does possess a strong odor) and should be added directly to the crystallization
drop when possible. Another consideration when working with DTT is to use the
microdialysis method for crystallization since DTT can be added to the dialysis
solution to replenish the supply of reducing agent.
TCEP HCl is stronger than both beta-me and DTT, lasting about 2 to 3 weeks in a
typical crystallization experiment. TCEP hydrochloride can acidify the crystalliza-
tion solution. Like DTT, TCEP HCl needs to be added directly to the crystalliza-
tion drop, hence, microdialysis is a consideration.
Beta-me, DTT, and TCEP HCl can behave differently. Like all additives, if you find
a class of compounds has an effect on sample stability or crystallization, then
screen a variety of compounds in that class to see which one is best for your
application. What we are trying to suggest is that like all additives, one might
consider evaluating all three reducing agents to see which one works best for
your sample.
There have been reports where reducing agents have been used as successful
crystallization additives where there were no free sulfhydryls in the sample.
Reducing agents can bind metals and trace metal compounds, inactivating the
reducing agent and the metal. This can make heavy atom derivatization in the
presence of reducing agents, difficult and frustrating. EDTA can be added to the
crystallization experiment to avoid inactivation of the reducing agent by metals.
Keep in mind that if your sample needs metals for activity or stability that EDTA
will keep metals from your sample.
When working at an alkaline pH, beta-me and TCEP HCl are more stable than
DTT. TCEP HCl is more stable at acid pH than DTT.
DTT reduces nickel ions and can cause problems when purifying His-tagged pro-
teins. To avoid this complication, try beta-me or TCEP HCl as a reducing agent
instead of DTT when purifying His-tagged proteins.
L-cysteine is also a reducing agent. Its usefulness in crystallization is limited since
it likes to form small hexagonal plate shaped crystals. L-cysteine can be a useful
crystallization additive, but keep those pesky hexagonal plates in the back of your
mind when working with L-cysteine.
If oxidation of the sample is expected or anticipated, reducing agents should
be present during the preparation of the protein (when possible) and should
be included, added, or replenished during the final preparation of the sample
for crystallization. In vapor diffusion experiments, the reducing agent can be
included or added to the crystallization reagent in the reservoir to minimize or
prevent dilution of the reducing agent in the sample drop.
When deciding on the appropriate concentration of reducing agent for the
sample, consider the number of free sulfhydryls in the sample as well as sample
concentration. More free sulfhydryls and higher sample concentration mean that
one should consider using higher concentrations of reducing agent.
Arsenical compounds are not compatible with reducing agents. Cacodylic acid,
or sodium cacodylate is an arsenical compound and popular crystallization buffer
and is not compatible with reducing agents. Other compounds not compatible
with reducing agents include ammonium nitrate, hydroperoxide, potassium
perchlorate, sodium nitrate. Check the Material Safety Data Sheet (MSDS) for
these reagents for specifics about the incompatibilities of these chemicals with
reducing agents.
S is for Sample Preparation
Preparing the Protein for Crystallization
Lyophilization
Avoid lyophilization. Even though there are many examples of proteins which
crystallize after lyophilization (lysozyme, thaumatin, hemoglobin), lyophiliza-
tion is to be avoided when possible. If the protein is lyophilized, it needs to be
dialyzed before crystallization. Dialyze the protein against deionized water or a
stabilization buffer before crystallization. Dialysis will remove non-volatile buffers
and other chemicals which may have been present before lyophilization.
Ammonium Sulfate Precipitation
Avoid using ammonium sulfate precipitation as a final purification and/or
concentration step. It is often very difficult to completely remove all the ammo-
nium sulfate by a desalting column of dialysis. The remaining trace amounts of
ammonium sulfate can interfere with crystallization screening results and create
reproducibility problems. It is not uncommon for trace amounts of ammonium
Solutions for Crystal Growth
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sulfate in the sample to cause precipitation or excessive nucleation in screen
conditions containing polyethylene glycol and salt.
Batches
Avoid combining different purification batches for crystallization trials. Purification
conditions and procedures are never identical so each batch should be screened
separately.
Profile the Protein
Ideally, you will purify your own protein, but this is not always reality. So, it is
always a good idea to characterize your protein before beginning crystallization
experiments. Profiling your protein before crystallization can often provide valu-
able clues during screening and optimization of crystallization conditions. Assays
to seriously consider:
SDS-PAGE
Native PAGE or
Dynamic Light Scattering
IEF (Isoelectric Focusing) Gel
Mass Spectroscopy
The results of these assays can:
Determine the purity of the sample
Determine the homogeneity of the sample
Identify batch-to-batch variations
Identify stability problems with the sample
How Pure?
How pure should the protein sample be for crystallization trials? As pure as
possible. Thats some answer, is it not? Integrating common sense into the ques-
tion, we might arrive at the following answer. For initial screening, the sample
should be at least 90 to 95% pure on a Coomassie stained SDS-PAGE. Finally, it
does no harm to screen an impure sample as one can always perform further
purification. Remember, crystallization used to be considered a very powerful
purification tool (and still is!).
If the initial screen does not produce crystals, any promising results, or it
becomes next to impossible to improve crystal quality during optimization, one
should consider further purification of the sample.
Storing the Sample
Most proteins can be stored successfully at 4C or -70C.
Check with the person preparing the protein or compare your protein to a simi-
lar protein in the literature for best storage temperature.
Ideally, one should assay the activity and stability of the protein before storage and
then later on at various points in time to determine the sample storage stability.
Repeated freezing and thawing of the sample should be avoided. Aliquot the
sample into multiple small microcentrifuge tubes. Make the aliquots small enough
so that the entire aliquot can be consumed in the experiment after thawing.
Sometimes people like to add glycerol (10 to 50% v/v) to help proteins better
tolerate freezing. Avoid this if possible since it is often difficult to remove glyc-
erol by dialysis or filtration. The presence of glycerol is a crystallization variable.
Glycerol can behave as a precipitant, an additive, or cryoprotectant and therefore
can influence the outcome of a crystallization experiment.
In general, it is better to store proteins more concentrated than diluted. When
too dilute, adsorption of the protein onto the storage container can lead to
significant losses. However, precipitation can sometimes be a problem when the
protein is stored too concentrated.
Label samples clearly with the sample identification, batch identification, and
date of storage. Small cryo labels can be very useful here. Color coding samples
can be a nice organization tool. For the sake of easy organization and identi-
fication, it is sometimes more convenient to nest samples. For example, store
batches of small microcentrifuge tubes in 10 ml or 50 ml centrifuge tubes and
organize them by batch or sample.
Sample Handling
Be nice to your protein. Remember that proteins make an excellent food source
for microbes. Protect your sample from microbes by storing the protein at less
than 4 C and not leaving the sample for extended periods of time at room
temperature.
When thawing a sample or mixing a lyophilized sample into solution do not
shake or vortex the protein. Avoid foaming the sample. Foam can be a sign of
denaturation.
Allow the sample to equilibrate to the temperature where the crystallization
experiments will be set up and/or incubated before setting the experiment.
The field of crystal growth is full of opinions and controversy. There are several
opinions on what should be done with the sample just prior to setting the crystal-
lization experiment. Lets have a look at those opinions.
Some like to filter the sample through a 0.2 micron (or smaller, but be sure to
compare the MW of your sample to the pore size of your filter so as not to stick
your sample on the filter) pore size filter into a sterile container. Filtration can
remove microbial contamination (but not the proteases) as well as sample aggre-
gation. Turbid sample solutions with lots of precipitate should be solubilized or
centrifuged before filtration to avoid the ugly experience of sticking the sample
to a filter membrane. Use filters with the smallest possible dead volume to
minimize sample loss. Some of the centrifugal microfilration devices are certainly
worth consideration. Before filtering the sample, wash or flush the filter with a
small amount of the sample buffer / storage solution before filtering the sample.
This will test the filter for compatibility with your sample buffer and remove
any trace glycerol which can sometime be present from the manufacturer. If
possible, test filter a small aliquot of the sample, and measure the activity / OD
before filtering the entire sample. Do this to test the adherence of the sample
to the filter media. Read and follow the instructions supplied before introducing
the sample to the filter.
P A G E 8 - 1 4
cryst alli zat i on ti ps from a to z
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Some like to centrifuge the sample. Centrifugation removes large sample aggre-
gates and amorphous debris. Post centrifugation views can provide a visual clue
of aggregation/precipitate for seemingly clear solutions. Following centrifuga-
tion, use only the supernatant for crystallization trials.
Others prefer to avoid filtration or centrifugation before setting crystallization
experiments. One view is that the presence of amorphous material or aggregates
can enhance the changes for crystallization be acting as nucleants.
To Azide or Not
Sodium azide (NaN3) is an anti-microbial preservative that is sometimes used
to protect samples and crystallization reagents from microbial contamination.
Sodium azide is toxic and should be handled with care. Typical sodium azide
concentrations are 1 mM or if you prefer % measurements, between 0.02% and
0.1% w/v.
If you choose to use sodium azide remember that:
It is toxic to humans as well as microbes.
It is an inhibitor for some proteins and may become an unintentional ligand
for your sample.
It can interfere with heavy atom derivatization.
Some metal azides are explosive.
There are reports where eliminating sodium azide from the experiment
improved crystallization.
Alternatives to Sodium Azide Include Thymol and Thimerosal.
A final alternative to the use of antimicrobials is the use of proper sterile tech-
nique and materials. Sterile filter all samples and reagents into sterile containers.
Store samples and reagents at 4C or lower. Use sterile pipet tips. Keep your
work area clean. Develop a sterile technique with your crystallization setups.
With common sense, sterile reagents and sample, good technique, and sterile
pipet tips, one can successfully avoid the use of chemical antimicrobials in the
crystallization lab.
Record Keeping and Organization
It is prudent to write down and hold onto detailed notes concerning the puri-
fication, storage, and handling of the sample. It is obvious that one should also
maintain records of crystallization trials which should include:
Sample information
Name of sample
Sample identification (batch, storage location, storage temperature, etc)
Sample buffer composition, additives, ligands, etc.
Sample concentration
Crystallization experiment information
Method
Drop size and composition
Reagent composition
Temperature
Date
Person performing experiment
Although it may seem trivial, a little AR, and excessive, it is reasonable to write
down anything that could become a crystallization variable.
Maintaining accurate and complete records of experimental observations is obvi-
ously important and will be covered in more detail elsewhere. In general, try to
use both descriptive and quantitative comments when developing crystallization
records.
Maintain a clean and well organized workplace for your crystallization setups.
Cleanliness will help to prevent chemical, microbial, and miscellaneous contami-
nation. Organization will prevent errors and save time.
Questions to Ponder About the Sample
Does a similar sample exist and has it been crystallized?
Does the sample contain free cysteines?
Does the sample contain additives such as sodium azide, ligands, inhibitors,
or substrates?
Is the protein glycosylated?
Is the protein phosphorylated?
Is the protein N-terminal methylated?
At what temperature is the protein stable?
How does sample solubility and stability change with temperature?
How does sample solubility and stability change with pH?
Does the sample bind metals?
Is the protein sensitive to proteolysis?
What class of protein am I working with (antibody, virus, enzyme, membrane
protein)?
What have been the most successful approaches with my class of protein?
What is the source of the sample?
How was the sample purified and stored before it arrived into my hands?
What is in the sample container besides the sample (buffer, additives, etc.)?
Is the sample pooled purification aliquots or a single batch?
How much sample do I have and how much more is available?
How pure is the sample?
P A G E 9 - 1 4
Solutions for Crystal Growth
Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com 230 Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com
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P A G E 1 0 - 1 4
How homogeneous is the sample?
Does anyone possess any solubility information on this sample?
What is unique about this protein?
What is necessary chemically and physically to maintain a stable, active
sample?
T is for Temperature
Temperature can be a significant variable in biological macromolecule and
small molecule crystallization
1-5
. Temperature often influences nucleation and
crystal growth by manipulating the solubility and supersaturation of the sample.
Temperature has also been shown to be an important variable with phase separa-
tion in detergent solutions during membrane protein crystallization
7
.
Control and manipulation of temperature during the screening, optimization,
and production of crystals is a prerequisite for successful and reproducible crys-
tal growth of proteins with temperature dependent solubility. Christopher et al.
5
,
testing 30 randomly chosen proteins, found 86% demonstrated a temperature
dependent solubility and suggested that temperature induced crystallization
could be a generally useful technique. Temperature was shown to affect quan-
tity, size, and quality of the crystals, as well as sample solubility and preliminary
crystallization data.
One advantage of temperature is that it provides precise, quick, and reversible
control of relative supersaturation. Using temperature in addition to standard
crystallization variables such as sample concentration, reagent composition and
concentration, as well as pH, can increase the probability of producing crystals
as well as uncover new crystallization conditions for a sample. Additional crystal-
lization conditions may uncover reagent formulations more amicable to heavy
atom derivatization, cryoprotection, and optimization or at least offer options.
Temperature is amenable to control and can be used to carefully manipulate
crystal nucleation and growth. This control can also be used to etch or partially
dissolve, then grow back the crystal in an attempt to improve crystal size, mor-
phology, and quality or assist with seeding. Temperature control is noninvasive
and can manipulate sample solubility and crystallization without altering reagent
formulation.
Typically, crystallization screens and experiments are performed at room tem-
perature and possibly 4C. A reasonable range of temperature to screen and opti-
mize for protein crystallization is 4 to 45C and some proteins have been crystal-
lized at 60C (glucagon and choriomammotropin). Temperature incubations
above room temperature should be monitored closely for evaporation from the
drop and reservoir. A 2 l hanging drop vapor diffusion experiment at 37C can
evaporate in as little as 48 hours depending upon the plate and quality of seal.
Microbatch under paraffin oil can minimize evaporation problems. In the case
of room temperature incubations, temperature control and stability are often
minimal since the experiments may be left in the open room. In an open room,
temperature fluctuations may be significant, especially over a 24-hour period and
on weekends when thermostatic control of the room environment can fluctuate
10 or more. Incubation at 4C and other temperatures are often more stable
since the incubation is performed in some type of incubator. Another source of
temperature fluctuation occurs while viewing experiments. The light microscope
is a heat source and extended viewing can significantly alter the temperature of
small drops. Quick, efficient viewing can minimize temperature changes. Also,
remember to turn off the light source when leaving plates on the stage in one
position for more than a few seconds.
While controlled temperature can be important for consistent results, tempera-
ture fluctuation can be useful in obtaining high quality crystals by screening a
larger range of crystallization conditions
8
since for a sample with temperature
dependent solubility, changes in temperature can equate to changes in a crystal-
lization reagent condition. Hence, a sparse matrix screen takes on a new dimen-
sion when screened at multiple temperatures, or ramped over several different
temperatures over a period of time.
How does one test for the effect of temperature and temperature dependent
solubility without consuming a lot of sample? One solution is to set a single
crystallization screen at one temperature, allow the experiment to incubate for a
week, record the results, and then move the plate to another temperature. Allow
the experiment to incubate for a few days to a week at the new temperature and
record the results. If one notices change in solubility (i.e. clear drop turning to
precipitate, or precipitate turning to clear drops) between the two temperatures,
then the sample has temperature dependent solubility and temperature should
be explored as a crystallization variable.
Temperature gradients can be used for screening and optimization of proteins
with temperature dependent solubility. For screening, set the experiment at one
temperature, allow the experiment to equilibrate and then slowly change the
temperature to a second temperature. In general, ramp the temperature so that
the sample is exposed to an increase in relative supersaturation as the tempera-
ture changes over time. In other words, ramp from high to low temperature if the
sample is more soluble at high than low temperatures. A temperature gradient
or ramp allows one to slowly approach temperatures where a sample may have
a decrease in solubility with a corresponding increase in relative supersaturation.
Published examples of temperature gradient or temperature ramp crystallization
include elastase
9
(25 to 20C gradient), alpha-amylase
10
(25 to 12C gradient), and
insulin
11
(50 to 25C gradient).
To demonstrate how screening temperature could affect and enhance the results
obtained from a preliminary crystallization screen, the Hampton Research M6
Mini Temperature Incubator was used to screen 4 different temperatures. Using
glucose isomerase and Crystal Screen, sitting drop vapor diffusion experiments
were set using Cryschem plates at 4, 15, 25, and 37C. Drops were observed
daily and the results were quite interesting. Glucose isomerase crystallized in 19
conditions at 25C, 23 conditions at 15C, 28 conditions at 4C, and 12 condi-
tions at 37C. A similar approach with Trypsin, yielded crystals in 8 conditions
at 15C, 4 conditions at 25C, and 7 conditions at 32C. In the case of Trypsin,
a single set of Cryschem plates were set and the plates simply moved from one
temperature to another over a period of a week, scoring results before each
temperature change.
Temperature Tips
In high salt, proteins with normal solubility will be more soluble at cold than
at warm temperatures.
In low salt, proteins with normal solubility will be more soluble at warm than
at cold temperatures.
Proteins with normal solubility will precipitate or crystallize from a lower con-
centration of PEG, MPD, or organic solvent more slowly at low than at high
temperatures.
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P A G E 1 1 - 1 4
Diffusion rates are less and equilibration occurs more slowly at low than at
high temperatures. Crystallization may occur more slowly at low than at high
temperatures.
Temperature effects can be more pronounced at low ionic strength reagent
conditions.
Do not use the appearance or non-appearance of crystals at various tempera-
tures to gauge the effectiveness of temperature as a crystallization variable.
Rather, use the difference in the solubility at different temperatures to gauge
the effect temperature has on sample solubility. If an effect is observed,
explore temperature as a crystallization variable.
Temperature can affect different crystal forms and growth mechanisms
12
.
When incubating experiments below and above room temperature and view-
ing experiments at room temperature, condensation can be a problem. To
minimize and avoid condensation with vapor diffusion experiments, stack a
dummy plate with reservoir filled with water and sealed, at the bottom and
top of the stack of plates. This will slow the temperature change in the sand-
wiched plates and minimize condensation.
The microbatch method works well for temperature exploration. In a tra-
ditional microbatch experiment, the relative supersaturation of the system
does not change since, in theory there is no vapor diffusion. However, if the
sample exhibits temperature dependent solubility, temperature can be used
to manipulate sample solubility in a microbatch experiment. Another plus of
using microbatch is the lack of condensation while viewing the experiment.
Covers of microbatch plates can be removed for a clear view.
Condensation with a hanging drop can mean alteration of your drop when
the condensation mixes with the drop. Condensation with a sitting drop can
mean there will be no mixing of the condensation with your drop, unless the
condensation falls into the drop. Moral, sitting drop has less chance of mixing
with condensation.
To dry up condensation, add a small amount of concentrated salt solution to
the reservoir. Keep in mind this might also dry your drop a bit.
Ideally, one should set the experiment at the eventual incubation temperature
and all reagents, samples, and plates should be equilibrated to the incubation
temperature. This is a reality for room temperature and 4C setups for those
of us with cold rooms. For the rest of us, we can set the experiment at room
temp and then toss it into the incubator. Or, for 4C setups, one can cheat.
Simply incubate the reagents, sample, plates, and slides in the refrigerator
before setup. During the setup, place materials in a tray full of ice. Maintain
the plates on ice during the setup. Seal and move smartly to the 4C incuba-
tor.
Nucleic acid temperature stability allows one to examine temperatures
between 4 and 35C.
Increasing temperature increases the disorder of reagent molecules. Varying
the temperature of a crystallization experiment can manipulate sample-
sample as well as sample-reagent and reagent-reagent interactions. Such
manipulations may have an impact on interactions that control nucleation and
crystal growth. In addition, such interactions may have an impact on crystal
packing as well as the termination of crystal growth. Hence, temperature can
impact nucleation, growth, packing, and termination.
Temperature can be a habit modifier and change the crystal lattice. For exam-
ple, at temperatures below 25C and in the presence of sodium chloride and
acidic pH, the tetragonal form of lysozyme is favored. Under similar reagent
conditions above 25C, the orthorhombic form is favored
13
.
The preparation of heavy atom isomorphous derivatives can depend upon the
temperature of the experiment. In most cases, it seems the soak temperature
is the same as the crystallization temperature.
References
1. Giege, R., and Mikol, V., Trends in Biotechnology (1989) 7, 277.
2. McPherson, A., European J. Biochemistry (1990) 189, 1.
3. A. Ducruix and R. Giege, Editors, Crystallization of Nucleic Acids and Proteins: A Practical Approach,
IRL Press at Oxford University Press, 1991.
4. Lorber, B., and Giege, R., Journal of Crystal Growth (1992) 122, 168-175.
5. Christopher, G.K., Phipps, A.G., and Gray, R.J., Journal of Crystal Growth (1998) 191, 820-826.
6. Haser, R., et al., Journal of Crystal Growth (1992) 123, 109-120.
7. Garavito, R.M., and Picot, D., Journal of Crystal Growth (1991) 110, 89.
8. Drenth, J., Crystal Growth (1988) 90, 368.
9. Shotton, D.M., Hartley, B.S., Camerman, H., Hofmann, T., Nyborg, S.C., and Rao, L., Journal of Molecular Biology (1968)
32, 155-156.
10. McPherson, A., and Rich, A., Biochem. Biophys. Acta (1972) 285, 493-497.
11. T.L. Blundell, and L.N. Johnson, Protein Crystallography, Academic Press (New York) 1976, 59-82 (method by Guy
Dodson).
12. A. McPherson, Crystallization of Biological Macromolecules, Cold Spring Harbor Laboratory Press, 1999.
13. Ataka, M., and Tanaka, S., Biopolymers (1986) 25, 337.
U is for Units

Molarity is the number of moles of solute per liter and in represented as M,
such as 3.0 M ammonium sulfate. Molarity is the ratio between the moles of dis-
solved solute (solid stuff) and the volume of solution (liquid stuff) in liters. The
accepted volume of the solution is 1 L, so a 1M (molar) solution would be 1M =
1 mole of solute/1 L solution. Molarity is a way of determining the concentration
of a solution. Dilute solutions are typically expressed in terms of millimolarity
(mM) where 1 mM = 0.001 M. Typically, in crystallization we are asked to make
something like a 3.5 M solution of ammonium sulfate. To do this we need to
know the molecular weight of ammonium sulfate (132.14 g/mole), the volume
of solution to make (lets make 500 ml or 0.5 L), and the desired concentration
(3.5 M doh!). Then we calculate:
# grams required = (3.5 mole/liter)(0.5 liter)(132.14 g/mole) = 231.25 g
So, we then weigh 231.25 g of ammonium sulfate and add deionized water to
dissolve the ammonium sulfate and then adjust the final volume to 0.5 liter (500
ml). Do NOT add 500 ml of water to 231.25 g of ammonium sulfate. Ideally one
should use the most precise measuring instrument possible such as a volumetric
flask. The less desirable instrument would be a graduated cylinder and the least
desirable would be a beaker.
Molarity is typically used as a concentration unit for salts 1,6-hexanediol, deter-
gents, and some additives.
% w/v (percent weight/volume) is often used when formulating high molecular
weight polyethylene glycols which begin life as solids as well as some additives.
% w/v is the weight of a solute in a given volume. % w/v = gram per 100 ml. For
example, 50% w/v PEG 3350 is 50 g of PEG 3350 in a final volume of 100 ml and
Solutions for Crystal Growth
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P A G E 1 2 - 1 4
NOT 50 g of PEG 3350 plus 50 g of water. This means weigh 50 g of PEG into a
graduated cylinder or volumetric flask and bring the final volume to 100 ml with
water. To clarify, in order to make a 500 ml of a 30% w/v solution of PEG 8000,
simply weigh 150 g of PEG 8000 and bring it to a final volume of 500 ml in deionized
water. Often the mistake of adding 150 g of PEG 800 to 350 ml of water is thought
of as a 30% w/v solution. This is a huge no-no and makes one look like a lab rookie.
Get it right and look cool.
%v/v (percent volume/volume) is often used when formulating liquid, low
molecular weight polymers (PEG 400), organics (MPD), and organic solvents
(iso-propanol) into stock solutions. % v/v is the volume of a solute in a given
volume and NOT a volume plus a volume. For example, a 50% v/v iso-propanol is
50 ml of iso-propanol brought to a final volume of 100 ml with water and is NOT
50 ml of iso-propanol plus 50 ml of water. Once again, the latter is a big no-no
and makes you look like a Homer.
Additional comment regarding % w/v and % v/v. Do not ask someone What is
your definition of % w/v?. There is only one correct definition. Therefore all
other definitions are wrong. Keep life simple and use only one definition of %
w/v and % v/v. The correct definition. There is only one correct way to make
these solutions. There is one occasion where one can ask the question, What is
your definition of % w/v? and this is when someone new enters the lab and you
want to check them out and see if they know anything about growing crystals or
you want to steal your buddys crystallization reagents but you want to be sure
they make them properly before you risk getting into trouble for stealing junk
solutions.
% saturation is the concentration of material in solution as a percent of the
maximum concentration possible at the given temperature. A saturated solution
is one where there is equilibrium between undissolved solute and dissolved
solute. To make a saturated solution, a salt is added to water and often warmed
to enhance solubilization. Complete dissolution is desired. Upon cooling, some
of the solute (salt) will crystallize out and leave behind a saturated solution. The
actual concentration of a saturated stock depends upon the temperature of the
solution. For example, at 0C, 127.5 g of potassium iodide can be dissolved into
100 ml of water, but at 20C, 144 g of potassium iodide can be dissolved into 100
ml of water. Therefore, depending upon whether the solution is kept at room
temperature or in the cold, the concentration can be very different. % saturation
is an old school way to make salt solutions for crystallization. However, since we
often perform crystallization at different temperatures, the actual concentration
in the bottle, reservoir, or drop can be very different. Plus, exact reproduction of
a stock solution not only depends upon careful mass and volume measurement,
but also temperature. Keep life simple, avoid reproducibility hassles and stick
with M, % w/v and % v/v.
Mg/ml (mg/ml) is typically used to express or determine protein concentra-
tion. Oftentimes we like to whip up a batch of lysozyme to test a new or stupid
crystallization idea or simply prove to ourselves that we can crystallize a protein
(especially those of us working with kinases, membrane proteins, and other
chewing gum macromolecules). To make a 20 mg/ml lysozyme solution we
might weigh 20 mg of lysozyme and simply add 1 ml of buffer (20 mg plus 1 ml).
However, others might weigh 20 mg of lysozyme and add 980 l (0.98 ml) (20
mg in 1 ml). Same rough definition, but different concentrations of lysozyme.
Might not make a difference unless you are trying to reproduce someone elses
work and you want to be sure you are comparing apples to apples and not apples
to oranges.
A final note on units. Knowing how to properly calculate, formulate, and docu-
ment crystallization reagents is an excellent foundation for becoming a crystal
Jedi master. Just the same, knowing how to properly calibrate and use an analyti-
cal and top loading balance and select the appropriate glassware for the job are
often overlooked, yet important crystallization variables. Quality and consistency
are big time crystallization variables so it is important to learn and master good
lab technique. While nobody ever won a Nobel Prize for cooking, most Nobel
Laureates can cook up a storm.
V is for Vibration
Vibration is often associated (in a crystallographers mind) with excessive
nucleation. It is popular belief that the best crystals grow when there is NOT a
significant source of vibration. It is believed this is particularly important during
the optimization and production stages of crystallization. Hence, it is suggested
that during optimization and production, one set crystallization experiments and
avoid the burning curiosity to look at the plates every single day. Try leaving the
setups alone during optimization and production.
Since vibration can lead to excessive nucleation, it seems one might want to
move the plates some during screening. One typically does move plates dur-
ing screening as it is typically recommended to view experiments each day for
the first week and once a week thereafter until the drop dries out. Maybe we
need to build vibrators into our screening incubators. Along the same line, it
is sometimes observed that plates which have been sitting for some time, then
viewed, often have crystals forming in the days following the latent observation.
So perhaps one should consider giving the plates a Monty Python nudge-nudge
or move them about to stimulate nucleation.
To avoid vibration, place plates in cupboards, cabinets, or incubators that are
infrequently opened and closed. Installing hydraulic door dampers to prevent
slamming doors or cupboards is one way to reduce vibration. Some incubators
vibrate more than other units and peltier based heating and cooling incubators
may be desirable from this perspective. However, some of the circulation fans in
these units vibrate quite a bit, so be sure to check out the fan and even consider
replacing it with a higher quality unit with less oscillation and vibration.
W is for Water
The role of water in protein crystallization is significant as protein crystals are
highly hydrated. The amount of water in a drop or a crystal, the purity, the
chemical composition, and how the water interacts with itself, the sample,
and reagents are all critical variables in a protein crystallization experiment.
Consistency is typically a good thing to keep in mind when trying to reproduce
crystallization results. Being consistent with ones treatment, handling and use
of water can be a important crystallization. Consider the source of water. One
should use pure and fresh water for crystallization experiments. Type II analyti-
cal grade water with a resistivity of less than 18.2 M-cm, with a total organic car-
bon (TOC) of less than 30 ppb and low bacteria count is a good source of water
for protein crystallization. Purchasing and using a quality water purification
system is one part of the water source equation. Different water systems and
different methods of water purification produce different water. For example,
glass distilled water typically has a pH of 5.5 while reverse osmosis and deioniza-
tion systems typically produce water with a pH of 7.
cryst alli zat i on ti ps from a to z
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P A G E 1 3 - 1 4
Another part of the water equation is the proper use and maintenance of the
water purification system. Carefully follow the manufacturers use, maintenance,
and service guidelines. Establish regular quality control practices to ensure the
consistent quality of the water. Using a water purification cartridge beyond its
expiration date or failing to clean the system on a regular basis can lead to water
with higher resistivity as well as microbial contamination. Microbial contamina-
tion can change the pH of the water as well as expose your sample to proteloytic
modification. Use fresh water. Purified water is deionized and can leach zinc,
lead, copper, iron, aluminum and other substance from glass or plastic storage
containers. Purified water stored in plastic containers for several weeks has
nearly the same level of TOC as tap water.
Use fresh, pure water for your crystallization experiments. When having trouble
reproducing experiments, consider water, the difference in purity and the differ-
ence in source as potential crystallization variables.
X is for Xenon
Xenon is a noble gas that binds to specific sites in a macromolecule. Xenon-
protein complexes can often serve as heavy atom complexes for MIR structure
determination. Heavy atom derivatives of protein crystals can be produced by
pressurizing native crystals with xenon gas. In the vast majority of cases, the
modification of the mother liquor to determine soaking conditions is avoided
since the crystal and mother liquor are simply placed in a chamber pressurized
with xenon gas. Another advantage of working with xenon is that it interacts
weakly with a protein so isomorphism of the derivative with the native crystal is
high. Also, the number of binding sites as well as the binding occupancies can
often be changed by altering the pressure of the xenon gas. Xenon binding sites
often differ from heavy metal binding sites so it can be useful to try xenon when
traditional heavy atom soaks fail. Xenon binding is often reversible so if one has
very few crystals, the same crystal could be used for heavy atom soak.
The interaction between xenon and the protein is typically confined to weak dis-
persion forces and does not involve electrostatic interactions. Therefore, xenon
should bind to different locations of the protein compared to most other heavy
atom compounds. The weakness of the interaction between xenon and the
protein makes it unlikely that xenon will interfere with crystal contacts. Xenon
derivatives usually show good isomorphicity with the native crystal, resulting in
high phasing power.
Xenon seems to have little or no effect on the pH or ionic strength of the mother
liquor. The weakness of the interactions between xenon and proteins requires
a significant xenon concentration in the crystals mother liquor to enforce suf-
ficient occupation of a potential binding site so most uses of xenon reported in
the literature make use of high pressure equipment for xenon derivatization.
At this time is appears cryocooling does not alter the proteins xenon binding
properties.
Xenon derivatization can be performed at room temperature and data col-
lection with xenon derivatives can be performed at room temperature using
specially made pressure cells and capillaries or at cryogenic temperatures using
CryoLoops.
Another application of xenon in protein crystals is the crystallographic imaging of
disordered areas of lipids or detergents in crystals of membrane proteins.
References for using xenon as a derivative:
1. Binding of Xenon to Sperm Whale Myoglobin. Schoenborn B.P.; Watson, H.C.; Kendrew, J.C.
(1965). Nature, 207, 28-30.
2. Cavities in proteins: structure of a metmyoglobin-xenon complex solved to 1.9&Angring. Tilton,
R.F.; Kuntz, L.D.; Pesko, G.A. (1984) Biochemistry 23. 2849-2857.
3. Using Xenon as a Heavy Atom for Determining Phases in Sperm Whale Metmyoglobin. Vitali, J.;
Robbins, A.H.; Almo, S.C.; Tilton, R.F. (1991). Journal of Applied Crystallography, 24, 931-935.
4. On the Preparation and X-ray Data Collection of Isomorphous Xenon Derivatives. Schiltz, M.;
Prange, T.; Fourme, R. (1994). Journal of Applied Crystallography, 27, 950-960.
5. Successful flash-cooling of xenon-derivatised myoglobin crystals. Soltis, S.M.; Stowell, M.H.B.;
Wiener, M.C.; Philips, G.N.; Rees, D.C. (1997).Journal of Applied Crystallography, 30, 190-194.
6. Freeze-Trapping Isomorphous Xenon Derivatives of Protein Crystals. Sauer, O.; Schmidt, A.;
Kratky, C. (1997). Journal of Applied Crystallography, 30, 476-486.
7. Protein Crystallography at Ultra-Short Wavelengths: Feasibility Study of Anomalous Dispersion
Experiments at the Xenon K-Edge. Schiltz, M., Kvick, A., Svensson, O., Shepard, W., De
LaFortelle, E., Prange, T., Kahn, R. & Fourme, R. (1997). Journal of Synchrotron Radiation , 4,
287-297.
8. A method to stabilize reduced and/or gas treated protein crystals by flash cooling under a con-
trolled atmospher
9. Xavier Vermede et.al. J. App. Cryst, (1999) 32(3) 505-509
10. Better structures from better data through better methods: a review of developments in de novo
macromolecular phasing techniques and associated instrumentation at LURE. Fourme, R. et al
(1999) J. Synch. Rad. 6(4) 834-844
11. Timmins, P., Pebay-Peroula, E. & Welte, W. (1994). Biophys. Chem. 53, 27-36.
Y is for Yummy
Protein samples are yummy treats for microbes such as bacteria, yeast and fungus
which secrete proteases that like to chop your sample into tasty bites, ruining
your crystallization setup. These same microbial menaces also like to dine on
polymer based crystallization reagents such a polyethylene glycols and even buf-
fers. So even if the bugs and their secreted proteases do not chop your sample
into bits, they can degrade reagents, alter the pH of the solution, or generate
chemical species which can influence crystallization and even make reproducing
conditions a pain in the tush. To prevent or at least minimize your sample and
reagents from becoming Sunday Crystallization Brunch for microbes, consider
the following suggestions:
Sterile filter the sample into a sterile tube using a 0.2 micron filter before setup to
remove microbes. Do not leave the sample on the bench at room temperature
for extended periods. Store unused sample appropriately (4, -20, or -80C; only
you know best). Those of you working with engineers, physicists, computer
scientists, or folks from outside the life science field, take a moment to realize
that quarks, gluons, hard drives and other physical things these folks are used
to handling do not support microbial growth. Take a little time to help these
crystallization rookies understand what happens to samples left on the bench for
a day or three and the significance of keeping things clean to prevent microbial
(as well as chemical) contamination. Case in point Caller: My lysozyme stock
that was growing crystals no longer grows crystals. Tech Support: Where are
you storing the lysozyme when it is not being used? Caller: Same place as the
reagents. Tech: Which is? Caller: On the bench (i.e. room temperature).
Tech Support: For how long? Caller: Oh, not long, maybe a few days, a couple
weeks at the most. Hey, nothing wrong with these types of questions. We all
gotta learn some time. Just be sure and help that engineer, computer scientist
and new post doc from a physics lab who is helping you with your new AFM
machine or testing the high throughput robot, to understand proteins are food
for microbes. Who knows, maybe they will help you with the next Windows
update installation.
Solutions for Crystal Growth
Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com 234 Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com
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P A G E 1 4 - 1 4
Sterile filter water into a sterile container before formulating crystallization
reagents that cannot or should not be filtered (detergents, gels, and some high
molecular weight polymeric agents too big for filtering).
Sterile filter (0.2 micron pore size) buffers, salts, polymers, and diluted organics
into sterile containers. Sterile containers such as polypropylene or PETG plastic
are readily available today and are cost-effective, time-saving alternatives to glass
and autoclaving.
When using a stock that has been sitting about for more than a couple of weeks,
give the bottle a swirl and look for signs of microbial growth such as a settle, faint
white, off white or yellow to brown precipitate. Sometimes slightly precipitated
salts will resemble microbial growth to the untrained eye. To help differentiate
precipitate from microbial growth, try warming the solution in your hands for 5
to 10 minutes. If the material disappears it might well be precipitate. Microbial
growth will not dissolve. Although we are not endorsing the sniffing of reagents,
especially since some crystallization reagents can be hazardous, precipitates will
not smell, while microbial growth will often stink.
Use sterile pipet tips when pipetting reagents, sample, and water into plates.
Keep your filthy paws away from your pipet tips and do not touch pipet tips to
the counter, your lab partner or other non-sterile items. If you set your pipet
down it had better not have a tip on it or the tip will likely touch something
non-sterile.
Keep crystallization plates in their sealed wrappers until just before use. This
will prevent airborne microbes from setting up home in your crystallization
plates. Most crystallization plates are not offered with a claim of sterility but most
makers of these plates operate the injection molds in clean areas and package
the plates straight out of the mold so there is little risk of contamination. Use
them as is and put the worry into keeping them clean once the plate is out of
the package.
If you do crystallization in a lab that also grows microbe bugs, especially fungal
microbes, be especially careful as these airborne spore forming nasties can set
up camp in crystallization plates quite easily.
Z is for Zeppenzauer
Microvolume crystallization by dialysis was first reported by Zeppenzauer in 1971
(Methods in Enzymology, ibid ref. 1. Vol. 22, page 253). The Zeppenzauer cells
are homemade from capillaries with the ends of the tubes either covered with
dialysis membrane or plugged with acrylamide, silica, or agarose gel. If cover-
ing the tube with a dialysis membrane, one can use small o-rings to secure the
membrane over the tube. The capillary is then placed in an appropriate chamber
with a volume of reagent sufficient to cover the ends of the tube.
Z is for Zinc
1 to 5 mM zinc (most frequently as zinc chloride or zinc sulfate) can be a useful
crystallization additive and typically reduces the solubility of the sample. Zinc
is not very soluble and can readily precipitate or crystallize out of reagents as
higher concentrations.

Z is for Zero
Zero or 0 is often used to represent a clear drop when scoring crystallization
experiments. While on the subject of zero or nothing, consider using -1 or some
other more interesting and exciting score to represent a non-experiment. A non-
experiment is a drop that has fallen off or a drop that was never pipetted (human
or robot error). Better to use a non-experiment score such as -1 than a 0 since
someone reading your scores might mistake a 0 as a clear drop.
cryst alli zat i on ti ps from a to z
1991-2009 Hampton Research Corp. All rights reserved. Printed in the United States of America.
This guide or parts thereof may not be reproduced in any form without the written permission of the publishers.
crystalli zati on ti ps
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Additives
Glycerol
Looking for yet another additive to try during optimization? Try Glycerol. Typical concentrations in the drop can range from 3 to 15%. Glycerol may limit
non-specifc aggregation and could be useful if you plan to use cryo techniques during data collection.
What Additive to Use
When trying to decide which additive might be useful during crystallization, try the following. If one has crystals and wants to try using additives to improve
the crystal size or quality, go back to the initial crystallization screening plates. Review the plates, looking for conditions where neither precipitate nor crys-
tals were/are observed. Now, review the results, looking for a common reagent ingredient present where no precipitate or crystals are found. For example,
one might fnd that all drops with Isopropanol remained clear. One could then try adding Isopropanol to the current crystallization condition to see if it
could improve the crystal size and/or quality. If one observes a difference in the crystal in the presence of isopropanol, then one might consider evaluating
other additives in the class of alcohols such as Ethanol, Methanol, tert-Butanol and others. If one has no crystals, but plenty of precipitate, phase separa-
tion and clear drops, follow the above analysis and try adding the common reagent ingredient found in the clear drops to those drops which contained
precipitate or phase separation. It is possible this agent could improve or at least manipulate sample solubility. The above tip was submitted from Jarmila
Jancarik from the laboratory of Sung Ho Kim at the University of California, Berkeley. Thank you Jaru! When using this approach it might be reasonable to
discern between concentration independent and dependent precipitation when trying to decide which agent to pursue as an additive. Try evaluating the
concentration independent agents frst and then look at the other agents if sample quantity permits. For example, if one observes precipitate in 15 to 40%
PEG but not in 5 to 10% PEG, it might simply be a concentration. However, if one observes no precipitate in 45 to 60% v/v MPD, one could guess that MPD
is a reasonable agent to evaluate as an additive.
Crystal Screen as an Additive Screen
Although Hampton Research does offer a specifcally formulated Additive Screen (HR2-428), here is a tip submitted by the crystallography group at
GSK (North Carolina) when one already has a sparse matrix screen or two laying about the lab. When screening additives, try adding 50 l of each Crystal
Screen reagent to 950 l of the "best" crystallization conditions thus far, in order to see if any of the reagents in Crystal Screen might serve as good addi-
tives. Crystal Screen 2 is an especially good kit for this technique since it contains numerous divalent cations such as Jeffamine

Reagent and a few other

"novel" agents. Jeffamine

is a registered trademark of the Huntsman Petrochemical Corporation.

Solutions for Crystal Growth
P A G E 1 - 2 9
Benzalkonium Chloride
Try Benzalkonium chloride (Fluka 12060). This cationic surface active agent has been reported to be useful as a crystallization additive with membrane
proteins and may be useful for soluble proteins. We've been using it in the drop between 1 and 3% w/v in water. Try a 10% w/v stock solution in water and
dilute into the drop to 1-3%.
Sodium Thiosulfate to Prevent Intermolecular Disulde Bridges
The presence of Thiosulfate in the protein solution was essential to promote crystal growth and to avoid the formation of unstable and weakly diffracting
crystals.
1
This is likely to be a consequence of the intrinsic capability of the reduced thiol group of the active-site cysteine to form disulfde bridges, leading
to the destabilization of the protein native structure. Sulfane sulfur-donor compounds such as Na
2
S
2
O
3
are likely to either keep the protein in the persulfu-
rated form or to prevent intermolecular disulfde bridges leading to unfolding and aggregation.
2
References
1. Crystallization and preliminary crystallographic characterization of LmACR2, an arsenate/antimonate reductase from Leishmania major. D. Bisacchi, Y. Zhou, B. P. Rosen, R. Mukhopadhyay and D. Bordo. Acta Cryst. (2006). F62, 976-979.
2. Bordo, D., Forlani, F., Spallarossa, A., Colnaghi, R., Carpen, A., Bolognesi, M. & Pagani, S. (2001). Biol. Chem. 382, 12451252.
Dissolving Hydrophobic Additives into Oil
Try dissolving the small molecule additive into paraffn or silicon oil, and use this mixture to cover the sample drop. This can be used with sitting drop, vapor
diffusion, or with microbatch under oil methods. The oil acts as a reservoir that may contain excess small molecules that (you hope) will be fed into the crystals.
cryst alli zati on ti ps
Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com 236 Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com
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Aggregation
Cryo
Sample Buffer
Non-specifc aggregation problems? Watch your sample buffer. The selection of an appropriate sample buffer/pH can often ward off aggregation problems.
Cryo Buffer
We've had good success using the well solution directly as the foundation of a cryobuffer in several situations where crystals cannot be grown directly in
the presence of cryoprotectant, and where crystals don't tolerate transfer to artifcial mother liquors. The basic protocol is as follows: (1) Remove 100 l of
the well solution after crystals have grown; (2) Split this sample into two 50 l aliquots; (3) Add 7.5 mg of Dextrose (Glucose) to the frst aliquot and 15 mg
to the second aliquot. Dissolve by gentle pipeting with a wide-bore tip. This will give two sequential well solutions that now contain 15% and 30% w/v Dex-
trose. If all the Dextrose won't go into the second aliquot, spin hard and remove the supernatant; (4) Transfer the crystal to aliquot number 1, equilibrate
for 3 minutes, then to aliquot number 2, then freeze. We've had a few crystals that routinely crack or blow up when transfered to artifcial mother liquor that
behave well when transfered to well solution plus Glucose. We assume that there is some aspect of the crystal drop (pH, ionic tension, precipitant concen-
tration) that is more effectively reproduced within the well than by separately prepared mother liquors. The nice thing about the protocol above is that you
don't get much of a volume increase when dry Dextrose is dissolved in the well solution, so the components in the solution are not diluted. Finally, if you
don't get a really good freeze, you can try to add about 5% v/v Glycerol to aliquot number 2, in addition to the 30% w/v Dextrose.
PEGs for Cryo
High molecular PEGs are also good cryoprotectants. If crystals are obtained from relatively high concentration of PEGs (e.g., 30% of PEG 3350), you can
cryoprotect them simply by raising the concentration of the PEG a little bit (e.g., 40% of PEG 3350).
J. Appl. Cryst. (2006). 39, 244-251
Effects of cryoprotectant concentration and cooling rate on vitrifcation of aqueous solutions. V. Berejnov, N. S. Husseini, O. A. Alsaied and R. E. Thorne
Synopsis: Critical concentrations required for vitrifcation of aqueous solutions are determined for fourteen common cryoprotectants, for sample volumes
ranging over four orders of magnitude and covering the range of interest in protein crystallography.
Mounting Thin Crystals
To mount very thin crystals onto CryoLoops

, frst dip the nylon loop into 0.5% Formvar

solution (Fluka # 09819) to form a thin flm. The flm provides

extra support for fragile crystals, and can result in much sharper refections with just slightly higher background. To clean the loop, dip it in alcohol to dis-
solve the support. Two notes: (1) the technique works only for crystals grown without organic solvents; and (2) take precautions not to breathe vapor from
the Formvar solution--the solvent is 1,2-dichloroethane. It is a standard support for electron microscopy grids.
The Potential Benets of Cryogenic Data Collection
Reduced radiation damage.
Decreased thermal motion and disorder.
Potential for improved resolution.
Increased crystal lifetime.
Crystals can be stored and shipped.
P A G E 2 - 2 9
P A G E 3 - 4 7
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Cryo continued...
Cannot Get Cryo to Work?
Try x-ray data collection at room temperature.
Evaluate other cryoprotectants. Try CryoPro

from Hampton Research (HR2-132).

Try the idential procedure again with another crystal.
Vary the time and temperature of the crystal handling steps.
Try annealing.
Match the osmotic pressure of your cryoprotectant to the osmotic pressure of the reagent producing the crystal. Crystallization reagents with lower salt con-
centrations require a higher percentage of cryo reagent for cryo protection than crystallization reagents with higher salt solutions (Garman 1999). Osmolality
tables (Weast 1988-1989) can be used to estimate the osmolality of reagents.
References
1. Cool data: quantity and quality. Elspeth Garman. Acta Cryst. (1999). D55, 1641-1653.
2. Weast, R. C. (1988-1989). Editor. Handbook of Chemistry & Physics, 69th ed. Boca Raton, Florida: CRC Press.
Cryo and Detergents
Be aware of the potential for detergent concentration mismatch between your mother liquor and the cryosolution. This particularly happens with vapor
diffusion setups: there is a delicate balance of "free" detergent in the mother liquor versus the proportion of the detergent which is bound to the protein.
Dropping a xtal into the cryosolution shocks the crystal with a bolus of extra free detergent. Hence, and counterintuitively, you may need to reduce the
detergent concentration in the cryosolution to keep everything in balance. Try titrating down from 1% to even as low as 0.4% in the cryosolution. Under the
conditions you are using, the CMC of bOG is suppressed below the usual 0.67% (w/v).
Also, the behavior of many of the alkyl glycoside detergents is very temperature sensitive. So be careful about the temperature of all the solutions you use.
R. Michael Garavito, Ph.D.
Submitted to CCP4 bulletin board February 2007
Edited by Hampton Research Corp.
Cleaning a CryoLoop

Soak the CryoLoop in a 0.5-2.0% detergent solution for 15 minutes or more. To improve the action of the cleaning solution, try increasing the temperature
of the solution. The use of ultrasonic or other agitation will further improve the cleaning action. Finally, lengthen the dwell time or increase the concentra-
tion of the solution.
Paper Wicks

can be used for delicate scrubbing of the CryoLoop. Dip the end of the wick into a detergent solution or 2-Propanol, and gently clean the
nylon CryoLoop.
Compressed air can be used to remove debris from a CryoLoop. Hold the tip of the air sprayer 3 inches from the CryoLoop and use gentle
pressure (less than 25 psi).
Paper Wicks - 55 mm X Fine Long - 100 wicks (HR4-211)
Paper Wicks - 55 mm Medium Long - 100 wicks (HR4-213)
Duster - Canned Air - 10 oz can (HR4-411)
Micro-90 Concentrated Alkaline Cleaning Solution (International Products Corporation, www.ipcol.com, 609-386-8770)
Solutions for Crystal Growth
P A G E 3 - 2 9
Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com 238 Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com
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Detergents
Detergent to Protein Ratio and Pea Type as Crystallization Variables
Ben-Shem et al (Acta Cryst. (2003). D59, 1824-1827) in the crystallization of higher plant photosystem I found the detergent to chlorophyll ratio had to be
carefully optimized in all purifcation steps in order to produce ordered crystals. Crystals were produced in 22.5 mM MES-bis-tris pH 6.6, 0.5% v/v PEG 400,
8.1 mM Ammonium citrate, 6% w/v PEG 6,000. Crystals appeared in 2 to 3 days and matured in size within two weeks time, yet the loss of sharp edges and
a degradation of diffraction quality was observed after an additional two weeks time. The initial diffraction resolution of 20 Angstrom was improved to 6
Angstrom through a seemingly tedious refnement of isolation, crystallization and cryo conditions. Again, the detergent to chlorophyll ratio as well as the
pea type and growing conditions with adjustment of the preparation to seasonal changes were essential to improvement of crystal quality.
Amphophiles
When screening detergents as additives, be sure to evaluate small amphophiles such as 1,2,3-Heptanetriol, Benzamidine, Ethanol, Dioxane, 1,6-Hexanediol,
Ethylene glycol, and Butyl ether for their ability to "manipulate" micelles.
Detergents, Oils and Microbatch
Detergents can be used successfuly as crystallization reagents in oil based microbatch experiments. Measurements by Barenda et al and Loll et al indicated
no signifcant loss of detergent will occur by migration of the detergent into the oil.
References
1. Thomas R.M. Barenda and Bauke W. Diskstra. Oils used in microbatch crystallization do not remove a detergent from the drops they cover. Acta Cryst. (2003). D59, 2345-2347.
2. Loll et al. Compatibility of detergents with the microbatch-under-oil crystallization method. Acta Cryst. (2003). D59, 114-116.
Handling Crystals
Separating Twins
The following procedure for separating twins is reported in "Structural Basis for Double-Stranded RNA Processing by Dicer", Ian J. MacRae, Kaihong Zhou,
Fei Li, Adrian Repic, Angela N. Brooks, W. Zacheus Cande, Paul D. Adams, Jennifer A. Doudna. Science 13 January 2006: Vol. 311. no. 5758, pp. 195 - 198.
Crystals of the protein Dicer grown at 18C by vapor diffusion in hanging drops composed of equal volumes of protein solution (9 mg/ml) and reservoir
solution (28% PEG-400, 0.1 M MgCl
2
, 0.1 M NaCl, 5 mM TCEP, 1 mM DTT and 0.1 M MES, pH 6.5). Resulting crystals were pseudo-merhodrally twinned,
producing an apparent spacegroup of P4222. Substituting MgCl
2
with MnCl
2
resulted in crystals that were macroscopically twinned. Individual crystals,
which belong to the spacegroup P21212, were extracted by soaking the largest twinned crystals in reservoir solution containing 15% PEG-400 to weaken the
twinning lattice contacts, followed by gentle mechanical prodding with a cat whisker. The resulting crystal shards were transferred back into full strength
reservoir solution, cooled to 4C for 1 hour and then cryo-cooled by plunging into liquid N2.
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P A G E 5 - 2 9
Handling Crystals continued...
14 Things to Say About Twins
1. Try additives.
2. Try DNA shuffing to introduce random mutations.
3. Try agarose gel crystallisation.
4. Try a new construct ("having tried everything else for 3 years before...").
5. Try to work on very small crystals and/or using a very small beam of a microfocus beamline at a synchrotron to isolate a single domain
and get less twinning.
6. Destabilise the crystal to separate the two halves (when possible) as in http://www.sciencemag.org/cgi/data/311/5758/195/DC1/1
7. If the crystals are obtained with Mg exchange it to Mn.
8. Change in crystallisation pH.
9. Change in crystallisation temperature.
10. You can try switching proteins. Lysozyme usually does the trick.
11. Co-crystallisation with partner proteins/domains or with ligands.
12. "Hmmm. Who knows, eh?"
13. Change crystallisation setup (e.g., from microbatch to hanging drops).
14. Test crystals at room temperature as well: "(EBV protease) which became twins only upon freezing, at room temperature the crystals were untwinned
(but resolution was much worse as well)".
Source: CCP4 Newsgroup, Stefano Benini Ph.D.
Crystal Annealing
Macromolecular crystal annealing (MCA) can help overcome increase mosaicity associated with cryocrystallography (Harp et al 1998, 1999). One might
consider annealing if diffraction is uncharacteristically poor after fash cooling. The process cycles a fash cooled crystal to ambient temperature and then to
cryogenic temperature and requires no special equipment. The annealing process does not improve a poorly diffracting crystal suffering from molecular dis-
order. The annealing protocol assumes that adequate cryo protection is available or that the crystal may be fash cooled using an oil and that the crystal dif-
fracts well. The crystal is fash cooled a cryo stream. In the MCA procedure, the crystal is removed from the cryo stream and placed in a large (300 microliter)
volume of the optimal reagent/cryo/oil solution from which the crystal was originally grown/mounted. Cover this drop to prevent evaporation. Incubation
time at room temperature is typically 3 minutes. Extended incubation times are okay, but shortening the incubation time can produce inconsistent results.
MCA has been successfully reported for both small and large crystals as well as for crystals with low (30%) or high solvent (70%) content.
Other forms of crystal annealing, one termed fash annealing (Yeh and Hol 1998), the other termed annealing on the loop (Harp 1999), have been used
successfully, especially on crystals with low (30%) solvent content. For fash annealing the crystals remain in the loop and on the mount. The cryo stream is
diverted for 1.5-2.0 seconds, then refash cooled for 6 seconds before repeating the process for a total of 3 rounds of rewarming and fash cooling. For an-
nealing on the loop, a variable length of time for warming is used without multiple rounds of warming and refash cooling. Warming time for cooling on the
loop is determined by observing the crystal in the loop while the stream is diverted and waiting until the drop in the loop is clear before refash cooling
the crystal. Warming time is typically proportional to the size of the crystal. For the fash annealing and annealing on the loop methods one might carefully
reduce the amount of liquid in the loop using a paper wick or micro wick as a variable to improve annealing results. But be careful not to let the drop dry too
much. Finally, one might be prepared for the formation of ice on the crystal and loop when the stream is blocked. Ice can be removed with the delicate use
of a paper wick or fber.
In closing, one may attempt the quick annealing methods frst, although the MCA seems to be more general.
References
1. Macromolecular crystal annealing: evaluation of techniques and variables. Harp et al. Acta Cryst. (1999). D55, 1329-1334.
2. Macromolecular crystal annealing: overcoming increased mosaicity associated with cryocrystallography. Harp et al. Acta Cryst. (1998). D54, 622-628.
3. A ash-annealing technique to improve diffraction limits and lower mosaicity in crystals of glycerol kinase. Yeh and Hol. Acta Cryst. (1998). D54, 479-480.

Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com 240 Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com
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Handling Crystals continued...
Minimizing Radiation Damage
Small molecules which behave as electron and free radical scavengers can be used for co crystallization and soaking to reduce radiation damage to the
macromolecular crystal. Recommended scavengers include oxidized glutathione, nicotinic acid, 5,5'-Dithiobis(2-nitrobenzoic acid) (DTNB), and ascorbic
acid. Typical scavenger concentration is 0.2 M in the original crystallization reagent. A quick (less than 20 second) soak can be used to introduce the
scavenger to the crystal. Prepare fresh solutions (add solid material to crystallization reagent in plate well) just before the soak.
References
1. How to avoid premature decay of your macromolecular crystal: a quick soak for long life. Brice Kauffmann, Manfreid Weiss, Victor Lamzin, and Andrea Schmidt. Structure 14, 1099-1105 July 2006.
2. Investigation of possible free radical scavengers and metrics for radiation damage in protein cryo-crystallography. Murray and Garman. J. Synchrotron Radiation 9, 347-354, 2002.
3. Blundell, T.L. and John, L.N. (1976) Protein Crystallography (New York, Academic Press).
Microbatch Crystal Mounting Tips
Here is the compiled 'tricks of the trade' for microbatch crystal mounting from the CCP4B community. Rebecca Page, Brown University, September 2006.
Original Post to CCP4 community:
I fnd mounting crystals grown in microbatch drops diffcult compared to mounting crystals grown out of sitting drops, primarily because of the oil layer.
Do most people simply use the oil layer as a cryoprotectant? If not, do you typically try to remove the oil prior to transferring the crystal to a cryoprotectant?
I'd like to compile 'tricks of the trade' for mounting crystals out of microbatch plates. Any advice would be helpful and I'll compile the list of responses and
repost for the community.
Summary of responses:
Sorry if this sounds like a crazy suggestion, but sometimes the simplest things work. Did you try to mount the crystal directly on a loop and see if it diffracts?
The oil can be cryoprotectant.
The setup would be:
1. Fish the crystal with a loop
2. Do not care if it comes across the oil layer and it retains the oil
3. Mount in the cryostream
4. Shoot x-rays and see if you have diffraction
------------------
If everything fails and you have few more drops of crystals and do not know how to freeze mount, here is another way you can try :- If you have few crystals
in the drop under oil and the drop size (excluding oil!!) is few l: add 10 l of mother liquor (you can get the mother liquor conc. based on few trials; it
should be few % more than the fnal conc. of the one you had used while setting up the microbatch, a decent start will be 5 % more ), allow it to stand for
few minutes. Then use the classical cappillary mounting method to suck the crystals slowly out and onto a cover slip. If possible try to remove as much as
possible (you may not be able to remove everything) the halo of oil surrounding the drop on cover slip (using the same capillary watching under micro-
scope to make sure you are not sucking out the crystals). The use of mother liquor (the 10 ul) is to basically to reduce the oil that come along when you
suck the crystals, so if necessary you can use more l's. Now quickly scoop the crystal with a loop and dip in the cryoprotectant and freeze it, in the usual
way. This has worked for me, though it may need some more standardization depending on your case.
------------------
In one case I had crystals grown under paraffn oil in microbatch, which required 25% glycerol for cryoprotection. However, when I transferred them directly
to the base condition + 25% glycerol, they would invariably break up. So I ended up picking up a crystal with a loop, pulling it through the oil, and putting it
on a coverslip. Then I removed the surrounding liquid and quickly replaced it with 4 l of base solution + 5% glycerol, observing the process under the mi-
croscope. The crystal seemed to hold up fne, and I then progressively replaced the solution with base condition + 10%, 15%, 20% and fnally 25% glycerol.
The resulting crystal froze nicely, and provided a 2A dataset. Any oil that was still around the crystal from the frst transfer was probably completely removed
by the cryo/washing steps, as other people have already described.
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Handling Crystals continued...
Solutions for Crystal Growth
P A G E 7 - 2 9
Microbatch Crystal Mounting Tips...continued
We do the same, I prepare 2-3 drops of cryoprotectant on a microscope slide, draw the crystal through the oil and "wash" in successive cryo drops until I see
no more oil coming off. Paraffn oil itself is mot a good cryo protectant, if you want to use paratone oil you can use the same procedure, or frst get rid of the
paraffn oil in artifcial motherliquor drops, then tranfer to paratone.
------------------
I have very tough time mounting crystals from microbatch. Essentially, I was adding the cryoprotectant to the drop and then transferring crystals to another
drop of cryoprotectant. Yet, I lost many crystals before successful mounting. At times, I tried to remove the oil layer and then add cryoprotectant, but it was
always diffcult.
------------------
I harvested exclusively from microbatch under oil for a little over half a year due to a crystal form that grew in 20%+ volatile alcohol (hanging drop tended to
swirl about something ferce when unsealed). I found that the main problem is getting the crystal up through the oil, as paraffn oil tended to knock crystals
out of loops when pulling out of the aqueous layer. Al's oil is a little easier, but still not a cake walk. However, residual oil on the surface of the drop/loop
didn't signifcantly affect diffraction. It was a little harder to see the crystals in the loop with blobs of oil getting in the way, but not impossible
------------------
A few schemes that worked well for me:
1. Step-wise supplement the drop with small volumes of mother liquor + over-concentrated cryo. For example, if starting with a 2ul drop, add 0.5ul at a
time of mother liquor + 30-40% glycerol. Once you've done this 4-6 times, you'll have (relatively) gently moved into 20% glycerol while staying under oil.
Different crystal forms may demand different steps/concentrations of the cryo stock.
or...
2. Use a pipete to remove all but a thin layer of oil from the top of the drop and then increase the drop volume to 4-6ul with mother liquor. This leaves a
thin flm of oil to slow down evaporation, but isn't quite so hard to pull crystals through. If you aren't in a volatile condition, you could just remove all of
the oil by fooding the drop this way . Then transfer to a separate well with cryo (also covered with a slick of oil, if the condition is volatile).
3. As for crystal/loop handling, the hanging drop method of setting the loop parallel to the surface of the drop and then pulling up is awful for microbatch,
as this causes the oil to shove the crystal out of the loop almost every time. I found that 45 degrees to nearly perpendicular to the surface of the drop
tended to work best.
------------------
A nice way to use cryo is to add it under the oil into the drop let it sit for the time you need and just fsh it with the oil then the crystals will be already pro-
tected. It worked for me and others in the lab and it also enlarge the volume of the drop so you can easily fsh.
------------------
I don't typically use the oil as a cryoprotectant.
First I remove *most* of the oil from the plate, and then remove the bulk of the oil from the well where I'll be working. Next, I use a piece of clay to hold the
plate at an angle, which makes it easier to get at the well. Then, I either
(a) fsh the crystal directly from the well (through the remaining oil) and transfer it to a small drop of cryoprotectant on a cover slip; or
(b) I use a pipet to transfer the drop to a cover slip and then work from there (in the latter case, you'll bring some oil along, which will coat the drop after
you deposit it on the cover slip, which helps prevent evaporation and allows you to perform the subsequent steps at a leisurely rate).
Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com 242 Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com
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Handling Crystals continued...
Microbatch Crystal Mounting Tips...continued
We pour off the oil and then add a harvest solution (~10-20 ul to each terasaki plate well) with ~2% more PEG than that in the droplet (remember that the
drop concentrates with time - sometimes fne tuning of the harvest solution PEG concentration is required but generally 2% works.). This keeps the crystals
stable and displaces any oil remaining at the top of the droplet. Sometimes there is a skin on top of the droplet. This is removed by a capillary or acupunc-
ture needle. Then we remove the crystals as per sitting drop. We have tried to cryo-protect the crystals by just pulling them up through the oil but this has
not worked for our crystals. It may be possible to add a cryoprotectant to the harvest solution but we haven't tried it.
------------------
Comments
1. Most people simply leave the oil, and loop the crystal out through the oil.
2. The crystal must ft the loop well, or the surface tension will drag it off the loop.
3. People who are used to working with oil claim it is easier than sitting or hanging drop because the oil prevents evaporation.
Therefore you don't have to work fast.
4. A loop with a bent handle can be helpful, because you have to harvest crystals from above.
5. Tip from Jeroen Mesters: if you want to use the oil as a cryoprotectant, you must DRY the oil frst. Use ordinary paraffn oil, and put 1 ml aliquots into
Epi-tubes. Dry tubes e.g. in a SpeedVac overnight. Freeze tubes, and use for a day, then discard. To use as a cryoprotectant, place a drop of dry oil on a
glass slide. Drag the loop with the crystal through the oil along the surface of the glass. Small drops of mother liquor will stick to the slide. Remove as
much mother liquor from around the crystal as possible, and freeze in the normal way. (Have I got that right Jeroen?)
------------------
I've mounted many crystals from microbatch and sitting drop plates and haven't found it diffcult to mount xtals from either! (well, of course a bit diffcult
when I frst started mounting xtals!!)
Typically I try to remove the oil by dragging the xtals gently in the cryoprotectant drop until I see the oil layer is removed (at much as I can judge visually!).
It's a tricky job removing the oil completely, depending on the cryo (eg. paratone!!!!) so great attention is needed in doing this part! I transfer the xtals to a
fresh drop of cryo before mounting.
------------------
1. Try to change to hanging drop vapor diffusion, by halving the precipitant.
2. Flood the microbatch drop with the frst stabilizing/cryosolution, then remove the oil layer.
------------------
We usually dip the loop straight through the oil drop and try to 'fsh' out the crystal. We then transfer it to a fresh drop of cryosolution and scoop it back out
before mounting it in into the beam. This however isn't as easy as it sounds and takes a little practice. We do sometimes remove the oil drop (partially) to
make it a little easier. Furthermore, if we use oil as a cryoprotectant, we would transfer it to a paratone (not paraffn oil) drop before mounting the crystal.
I have never heard anything about using the paraffn oil on top of the drop as a cryoprotectant. However, I found out that using paratone as a cryoprotectant
is also very diffcult, and as for all the described techniques, practice makes perfect.
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P A G E 9 - 2 9
Crystal and Age
CryoLoops for X-Ray Data Collection at Room Temperature
How Does the Age of a Crystal Affect X-Ray Diffraction Data?
Reading and References:
The crystal structure of yeast phenylalanine tRNA at 2.0 resolution: cleavage by Mg2+ in 15-year old crystals. Luca Jovinea,, Snezana Djordjevica and Dan-
iela Rhodes. Journal of Molecular Biology, Volume 301, Issue 2, 11 August 2000, Pages 401-414.
Abstract: We have re-determined the crystal structure of yeast tRNAPhe to 2.0 resolution using 15 year old crystals. The accuracy of the new structure, due
both to higher resolution data and formerly unavailable refnement methods, consolidates the previous structural information, but also reveals novel details.
In particular, the water structure around the tightly bound Mg2+ is now clearly resolved, and hence provides more accurate information on the geometry
of the magnesium-binding sites and the role of water molecules in coordinating the metal ions to the tRNA. We have assigned a total of ten magnesium ions
and identifed a partly conserved geometry for high-affnity Mg2+ binding. In the electron density map there is also clear density for a spermine molecule
binding in the major groove of the T?C arm and also contacting a symmetry-related tRNA molecule. Interestingly, we have also found that two specifc
regions of the tRNA in the crystals are partially cleaved. The sites of hydrolysis are within the D and anticodon loops in the vicinity of Mg2+.
Atomic resolution structure of a succinimide intermediate in E.coli CheY. Simonovic M, Volz K. J Mol Biol. 2002 Sep 27;322(4):663-7.
Abstract: Isomerization of aspartate to isoaspartate occurs spontaneously in proteins, causes changes in protein structures, and correlates positively with
the aging processes of many organisms, including Alzheimer disease in humans. Aspartate isomerization proceeds through an unstable cyclic succinimide
intermediate. There are few protein structure determinations that have characterized the intermediates and products of this isomerization reaction. Here we
report the discovery of an unusually stabilized succinimide ring in the 1.1A structure of the Escherichia coli CheY protein, as determined from a crystal eight
years old. The ring is formed by the side-chain of aspartate 75 and the backbone nitrogen of glycine 76 in an exposed loop of the molecule. Stabilization of
the succinimide is through interaction of a sulfate ion oxygen atom with the imide nitrogen atom. Formation of the ring caused conformational changes in
the loop, but did not alter the overall structure of the protein.
Using rational screening and electron microscopy to optimize the crystallization of succinate:ubiquinone oxidoreductase from Escherichia coli. R. Horse-
feld, V. Yankovskaya, S. Trnroth, C. Luna-Chavez, E. Stambouli, J. Barber, B. Byrne, G. Cecchini and S. Iwata. Acta Crystallographica Section D, Biological
Crystallography Volume 59, Part 3 (March 2003).
Synopsis: Crystals of SQR that diffract to 2.6 were obtained by rational screening and sample quality analysis using electron microscopy.
From the publication: "It proved critical to freeze the crystals within 72 h of crystallization set-up. Crystals that were frozen after this time limit showed no
diffraction. This alteration in properties was apparent by the change in colour from deep orange to pale yellow that was observed in crystals more than
four weeks old (Fig. 1c). The deep orange colour of the crystals is attributed to the presence of haem b within the protein. The loss or breakdown of haem,
demonstrated by the change of colour in the crystals, could lead to structural instability and consequently loss of diffraction. "
We use a much easier test for mounting crystals at room temperature. Just coat the crystal with Paratone-N (HR2-643) and mount your crystal in a standard
CryoLoop. The Paratone-N will slow down evaporation enough - no special tools required. You don't even need to remove all the liquid as you would do for
fash-cooling the crystal in Paratone-N. A major advantage is that you can use the same crystal to collect under cryo conditions and directly compare the impact
of cooling the crystal.
Reference: CCP4 Bulletin Board January 16, 2009 from Filip Van Petegem, The University of British Columbia.
Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com 244 Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com
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Methods, Procedures, Technical
Using Stock Solutions from the Refrigerator
When using stock solutions from the refrigerator or any one that has been standing for some time, be sure to thoroughly mix it before use. Storing stock
solutions in the refrigerator can result in condensation forming on the inside upper portion of the bottle. If this condensation falls into the stock solution, it
will create a heterogeneous stock concentration which in turn could make reproducing the experiment diffcult. Condensation can occur with any seal when
there is a temperature change in the environment. So it is possible for this phenomena to occur with room temperature stored stocks if the temperature in
the room changes signifcantly. To avoid such problems, it is generally good technique to mix crystallization reagents prior to pipetting.
Want Control and Accuracy?
Use a gel-loading pipet tip to dispense small drops with better control and accuracy of drop placement.
Mixing Drops
If you routinely mix the sample with the precipitant by aspirating and dispensing the drop several times, try setting up the drop without mixing. Simply
pipet the precipitant into the sample and let the two diffuse together without mixing. On the other hand, if you usually set up your drop without mixing, try
mixing. During the 2005 RAMC meeting, a show of hands survey among 100 participants showed most people do not mix their drops.
Make Connections
Communicate. Call (no e-mail allowed) a colleague and ask them their favorite crystallization trick or tip. Offer up one or two of yours.
PEG 400 and MPD
If you are using PEG 400 as a precipitant with some success, try MPD. If you are using MPD, try PEG 400.
Read the Classics
Read the classics. Try Laboratory Experiments in Biological Chemistry by James B. Sumner and G. Fred Somers, Academic Press 1949.
Trying Different Precipitants
Escape the local minima of using the same precipitants. If something worked once, great. Try it again, but remember to try something new.
Check Out Temperature
Try different temperatures. Room temperature and 4C are the norm and work quite fne. Anything between 0C and 37C is fair game. If you do not have
an incubator, simply look for a cool or warm space in the building to incubate the plates.
Free Interface Diffusion
Try free interface diffusion as a crystallization technique. This is easy to do and requires only a small amount of sample. Using a 20 l microcapillary pipet,
pipet 2 to 5 l of sample into one end of the capillary. Next, carefully pipet 2 to 5 l of precipitant SLOWLY into the capillary without introducing an air
bubble. Seal the ends with wax or clay. It helps to pipet the less dense material onto the material of higher density (i.e. pipet sample onto 30% PEG 8,000,
pipet 15% Isopropanol onto the sample). Use gel-loading pipet tips to load the sample and precipitant.
Removing Mother Liquor
Tired of cutting flter paper for wicks to remove mother liquor from capillaries during crystal mounting? Try our Paper Wicks

. The 55 mm X Fine Long

(HR4-211) work great in 0.75 mm capillaries.
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P A G E 1 1 - 2 9
Methods, Procedures, Technical continued...
PEG Sensitivity
Keep stocks and working solutions of PEGs in the dark. Use amber or solid colored bottles or wrap clear bottles in foil. If you want to be really careful, purge
with Argon and freeze them.
About Growth Rate
As a general rule, larger and fner crystals are obtained if the growth rate is minimized. If you are growing microcrystals, reducing their growth rate by more
gradual equilibration with the reservoir may result in their attaining larger size.
Reducing Agents
Consider using a reducing agent as a variable in the crystallization. Include a small amount (0.001 to 0.01 M) of a mild reductant such as TCEP hydrochloride,
Cysteine, B-Mercaptoethanol, Glutathione, or Dithiothreitol in the mother liquor. Besides protecting sensitive Cysteine residues, there is some indication
that antioxidants play some other undefned role in crystallization. Try levels of reducing agent much higher than that generally thought necessary to main-
tain sulfhydryl function.
Do Not Disturb
Once crystallization conditions are relatively well established, set up a crystallization trial and do not examine or disturb it in any way for several weeks.
Premature handling of crystallization trials can convert a few promising growth centers into a massive shower of microcrystals.
Using Sodium Azide
Keep crystallization samples and hardware free from microbial presence by fltering all solutions through a 0.22 micron flter. Store stocks and working
solutions in sterile containers. In most cases this negates the need for antimicrobials such as Sodium azide.
Bioseparation
Consider crystallization as a tool for bioseparation.
Temperature Effects
Proteins in high salt are usually more soluble at lower temperatures than higher temperatures. The reverse is generally true for PEG and MPD. Keep this in
mind during initial screening and optimization. Also keep in mind that this is not always true.
pH pHun
pH adjustments in the drop can be made by adding a small amount of concentrated Ammonium hydroxide to the drop or reservoir to increase the pH, or
Acetic acid to lower the pH.
Cold Organics
If you are using organic solvents as precipitating agents, be sure to include 4C incubations with your trials.
How to Dilute
A 1:10 dilution can be defned as 1 part of stock plus 9 parts of diluent. For example, to make 1,000 l of 0.1 M buffer from 1.0 M buffer, simply pipet 100 l
of 1.0 M buffer into a tube and add 900 l of diluent (water).
Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com 246 Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com
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Methods, Procedures, Technical continued...
What is Your Denition of % W/V?
Percent Weight/Volume (% w/v) = the weight in grams of a solute per 100 milliliter of a solution. For example, a 50% w/v solution of PEG 6,000 is made by
adding exactly 50 g of PEG 6,000 solid to a total, fnal volume of 100 ml. NOT by adding 50 g of PEG 6,000 to 100 ml of water.
Removing Urea and Acrylamide
Urea and Acrylamide contamination of RNA samples from denaturing gels cannot be removed by dialysis. Acrylamide and Urea can be removed using a
strong cation exchange resin. Joe Ng, UAB.
Path and Endpoint
The path is as important as the endpoint. Vary the method, drop ratios, and reservoir volume to alter the path to the same endpoint as a variable during
optimization.
Inorganic False Positives
Beware of inorganic salt crystals when using Phosphate, Carbonate, or Borate buffers!
Condensation
Tired of condensation problems in the plates at 4C? Fill two dummy plates with water, seal and place one at the top and the other at the bottom of your
stack of plates. Voila! No condensation problem!
Taking Care of PEGs
Polyethylene glycols are inherently unstable and are prone to produce aldehydes and peroxides. The presence of these oxidation byproducts increase
with the age of the PEG and is promoted in the presence of UV light. One can slow the production of aldehydes and peroxides by storing the PEG at low
temperatures and in the absence of light and oxygen. Flooding the storage bottles with Argon will help limit the oxidative effects of oxygen on the PEG. An
increase in the oxidative byproducts will inherently change the pH, conductivity, and ionic strength of the PEG.
Sodium Azide and Heavy Metals
Never use Sodium azide in crystallization reagents that contain heavy metals since the combination poses the danger of forming explosive metal azide salts
(See Problems associated with the use of azide as an inhibitor of microbial activity in soil, M. Rozycki and R. Bartha, 1981, Appl. Env. Microbiol. 41: 833-836).
Equilibration Kinetics
Recently we chatted about ways to alter the equilibration kinetics in a hanging or sitting drop vapor diffusion experiment. Another way to fool with the kinet-
ics is to perform the experiment at a cool temperature. Mikol et al have described how both the protein and precipitant are concentrated three to fve time
faster at 20C than at 3C. For details see V. Mikol, J.L. Rodeau, R. Giege, 1990, Anal. Biochem. 186, 332-339, Experimental determination of water equilibra-
tion rates in the hanging drop method of protein crystallization.
Physical Chemical Aspects
Always take into consideration the effect that salts, polymers, organic solvents, and additives might have on the pH of the buffer selected for the crystalliza-
tion experiment. To be safe, check the pH of an experimental solution after all of the reagent components have been added, mixed, and the solution has
equilibrated to the temperature where the buffer was titrated and the experiment is to be performed.
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Methods, Procedures, Technical continued...
Using Different Reservoir Volumes
Although one typically uses 1,000 l of reservoir solution in a VDX

or Linbro

Plate, one can actually get away with 500 l of reservoir solution before
having to begin worrying about the excessive effects of evaporation which occurs when using polystyrene crystallization plates. The utilization of less volume
in the reservoir can alter the kinetics of equilibration between the reservoir and the drop. Less volume equates to slower (longer) equilibration times. This
is not necessarily a bad thing when screening for preliminary crystallization conditions. For a good read on the kinetics of macromolecular crystallization see
"Kinetic Aspects of Macromolecular Crystallization" by Joe Luft and George DeTitta, page 110-130, Methods in Enzymology, Vol. 276,
Part A, 1997.
Crystal Annealing
Flash-cooling crystals can sometimes increase the mosaicity of biological macromolecular crystals. In some case, macromolecular crystal annealing can
reduce the mosaicity of fash-cooled crystals without affecting molecular structure. Crystal annealing involves cycling a fash-cooled crystal to room tempera-
ture and then back to cryogenic temperature. The procedure can also be applied to sometimes restore diffraction from fash-cooled crystals that were not
properly handled to and from cryogenic storage. Crystal annealing does not seem to improve a poorly diffracting crystal suffering from molecular disorder.
The essential features of the crystal annealing procedure are the following. The crystal is frst fash-cooled. The crystal is removed from the cryostream and
quickly transferred to a drop of cryoprotectant at crystal growth temperature and allowed to remain in the drop for at least three minutes. The drop should
be covered to prevent evaporation. Finally, the crystal is remounted on a loop and fash-cooled. Reference: Macromolecular Crystal Annealing: Overcoming
Increase Mosaicity Associated with Cryocrystallography, J.M. Harp, D.E. Timm, and G.J. Bunick, Acta Cryst. (1998) D54 622-628.
Isoelectric Homogeneity
Protein preparations which exhibit microheterogeneity are sometimes not amenable to crystallization or yield poorly diffracting crystals. Isoelectric hetero-
geneity is one such biochemical crystallization variable. Protein isoforms are often the result of modifcations such a phosphorylation, methionine oxidation,
or glycosylation. Such modifcations can cause very small changes in the isoelectric point (pI) of proteins. These isoforms can sometimes be resolved by
chromatographic techniques such as ion exchange (Crystallization of Isoelectrically Homogeneous Cholera Toxin, Westbrook & Spangler, Biochemistry
1989, 28 1333-1340). Conventional preparative isoelectric focusing techniques utilizing carrier ampholytes can often resolve isoforms, but suffer from pH
gradient instabilities and residual contamination of the proteins by ampholytes. An alternative technique that can avoid these complications, PrIME (Hoefer
Pharmacia Biotech) has been used successfully to further purify proteins which have subsequently been crystallized (Fab fragment of anti HIV gp-41 mono-
clonal antibody and isoforms of epidermal growth factor - Righettia et al).
Stabilizing Crystals with Glutaraldehyde
Soak crystals overnight in a solution comprised of the mother liquor utilized to grow the crystals plus 0.5 to 2% v/v Glutaraldehyde. Glutaraldehyde cross-
links proteins through the epsilon amino group of lysines. Cross-linking can be used to stabilize fragile crystals for seeding or data collection.
Reference: F.A Quiocho and F.M. Richards. Proc. Natl. Acad. Sci. U.S.A. 52, 833 (1964).
Building Blocks
During the growth of a high quality crystal, identical components are incorporated into a periodic lattice. Thus, sample homogeneity, the presence of
uniform biological macromolecules, is typically essential for the growth of high quality crystals.
Homogeneity
If members of a homogeneous population alter their form, then the population is no longer homogeneous. Stability of a population is important in order to
maintain homogeneity. One should take measures to preserve the stability of the sample and make it resistant to change, in order to prevent denaturation
and aggregation.
Read the Tea Leaves
We set screens in anticipation of growing crystals. In reality, we are looking for ways to alter the solubility of the sample in order to grow a crystal. Therefore,
review screens looking for more than crystals. Look for variables that alter the sample's solubility. The big hitters are sample concentration, reagent type and
concentration, pH, and temperature.
Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com 248 Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com
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Methods, Procedures, Technical continued...
Reverse Vapor Diffusion
Reverse vapor diffusion can sometimes be used to crystallize a protein in low ionic strength. This is an alternative method to dialysis that is less tedious
to set up but also allows the protein concentration to become more dilute as the experiment proceeds, whereas dialysis can maintain a constant protein
concentration. Begin with the concentrated protein in salt. The concentration of salt should be suffcient to maintain the solubility of the sample, perhaps
0.1 to 0.5 M salt. Set the sample for crystallization using the hanging or sitting drop vapor diffusion method. Pipet deionized water into the reagent reservoir.
Pipet the high ionic strength protein sample onto the cover slide (hanging drop) or post (sitting drop). Do not add water to the sample. Using only deion-
ized water in the reagent reservoir will allow water to diffuse from the reagent reservoir into the drop. As the drop size increases by vapor diffusion, the ionic
strength in the drop will decrease. The protein concentration will also decrease, so it is a good idea to begin with a concentrated protein sample. It is also
helpful to avoid having too much salt in the samples since this may cause the sample drop to absorb too much water and dilute the protein concentration.
For cryoprotection, one can add Glycerol to the reagent reservoir and mix this with the sample drop.
Gentle Crystal Cross-Linking with Gutaraldehyde
To cross-link fragile crystals for a chance at improved handling, solution changes or cryo, try the following: If you have hanging drops, then transfer the
cover slide to a Cryschem

24 well sitting drop plate or place a Micro-Bridge

into the 24 well hanging drop plate. Pipette 2 to 5 l of 25% Glutaraldehyde

into the sitting drop post and pipette your crystallization reagent into the surrounding reservoir. Seal the experiment. Allow the Glutaraldehyde to vapor dif-
fuse into the sample drop containing the crystal. Thirty minutes to six hours is generally suffcient. Glutaraldehyde relies primarily on lysine residues so the
number of lysines as well as temperature can be two signifcant variables infuencing cross-linking time. Note that amines will interfere with Glutaraldehyde
so Ammonium sulfate or Tris buffer will have to be dialyzed or exchanged out and substituted before cross-linking is attempted.
Note: Glutaraldehyde is toxic and precaution was taken against exposure to the reagent by conducting the cross-linking reactions in a chemical fume hood.
Reference:
1. J. Appl. Cryst. (1999). 32, 106-112 . A gentle vapor-diffusion technique for cross-linking of protein crystals for cryocrystallography. Carol J. Lusty
Cryschem Plate - Circle Your Hits
Circle your drops with crystals or promising results. With a Cryschem plate sealed using tape, simply use a Sharpie

to circle the drops that look promising

and those that have crystals. This makes it easy to spot these drops later during follow up. We suppose you could get fancy and use different colors to code
the results, but a simple black Sharpie circle around a well is easy to spot, even through a stack of plates.
Seeding from Wild Type Crystals to Grow Mutant Crystals
If you are able to obtain crystals of the native or wild type protein and cannot get crystals of variants or mutants of the native protein, try streak seeding.
Streak seed from the wild type crystals into drops containing the variant/mutant.
Glover et al. streak seeded from wild type crystals, either immediately or after a day of equilibration. The seeding stock solution was prepared by diluting
a drop with small crystals 100-fold using the reservoir solution. Streak seeding was performed by dipping a hair into the seeding stock solution and rinsed
twice in the reservoir. Crystals grew to 100-400 micron in 1 to 2 weeks.
Reference:
1. Principles of protein-DNA recognition revealed in the structural analysis of Ndt80-MSE DNA complexes. Jason S. Lamoureux and J.N. Mark Glover. Structure 14, 555-565, March 2006.
Using Evaporation to Control Crystal Nucleation
Jovine (2000) used an evaporation method to control excessive nucleation and produce larger crystals.
A vapor diffusion experiment is set with the sample and reagent concentration where no crystals form when the drop and reservoir are in equilibration with
one another. Nucleation is effected by opening each vapor diffusion experiment (remove cover slide, flm or tape) for a precise amount of time. The amount
of time used by Jovine was 100-120 seconds. The amount of time is empirically determined and will depend upon drop volume, reservoir volume, tempera-
ture, and reagent formulation. The opening increases the evaporation of water from the drop and creates a small and temporary increase in the relative
supersaturation, hopefully enough to promote nucleation before the system returns to equilibrium when the experiment is sealed. If the drop remains clear,
the procedure can be repeated until crystals start to appear. If too many crystals appear, reduce the amount of evaporation time.
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Methods, Procedures, Technical continued...
Using Evaporation to Control Crystal Nucleation continued...
This method can also be applied to experiments which remain clear, as a way to effect a change in the drop's relative supersaturation, which could result in
a crystal, precipitate or phase separation.
Reference:
1. A simple technique to control macromolecular crystal nucleation efciently using a standard vapour-diffusion set up. Luca Jovine. J. Appl. Cryst. Volume 33, Part 3, Number 2, 988-989 (2000).
Alternative Reservoirs in Vapor Diffusion Experiments
Setting up vapor diffusion crystallization experiments against different reservoir solutions may have a profound effect on the outcome of crystallization
experiments. It has been demonstrated that one way to increase crystallization space is to set the same conditions over different reservoirs (Newman 2005).
So, rather than screen more reagents, one can screen the same reagent set over two or three different reservoirs (such as Sodium chloride, Ammonium
sulfate and Polyethylene glycol 3,350), in addition to the crystallization reagent. Using the screen reagent in four sets of drops and then using the reagent in
addition to three alternative reservoirs expands the crystallization space as the path of equilibration and endpoint is unique for each reservoir solution.
Recommended reservoir solutions
1.25 - 2.0 M Sodium chloride
1.0 - 1.5 M Ammonium sulfate
30-50% PEG 3,350
20% PEG 3,350, 0.2 M Sodium chloride
Replacement reservoir solutions (approximate)
0.5 M Lithium chloride = 35% PEG 4,000
1.5-2.0 M Lithium chloride = 2.5 M Ammonium sulfate
Sodium chloride should be expected to act as a simple desiccant. Ammonium sulfate can desiccate and affect the pH of the drop through the gaseous NH
3

which is in equilibrium with the NH
4
+
in solution. PEG 3,350 should desiccate, although with different kinetics compared to the salts (Luft & DeTitta 1995).
Alternative reservoir solutions have also been referred to as common dehydrants, a generic reservoir, and desiccants. The use of a single, simple, concen-
trated solution for the reservoir in vapor diffusion crystallization experiments has been happening for decades (Dunlop & Hazes, 2005; Hempel, 1968; Luft
et al., 1994; McPherson, 1992).
To speed delivery of the reservoir for 48 and 96 well crystallization plates that are set up manually, the reservoir solution can be transferred to a Multichan-
nel Pipetter Basin (HR3-269) and delivered to the plate using a multichannel pipette. For automated experiments, the reservoir solution can be transferred
to a MASTERBLOCK

96 Deep Well Plate (HR3-105) or a Multichannel Pipetter Basin (HR3-269).

References :
1. Expanding screening space through the use of alternative reservoirs in vapor-diffusion experiments. Janet Newman. Acta Cryst. (2005). D61, 490-493.
2. Single Crystals of Transfer RNA from Formylmethionine and Phenyalanine Transfer RNA's.Arnold Hempel et al. Science, New Series, Vol. 162, No. 3860 (Dec. 20, 1968), 1384-1387.
3. A modied vapor-diffusion crystallization protocol that uses a common dehydrating agent. Dunlop & Hazes. Acta Cryst. (2005). D61, 1041-1048.
4. Luft et al. (1994). J. Appl. Cryst. 27, 443-452.
5. Luft & DeTitta. (1995). Acta Cryst. D51. 780-785.
6. McPherson. (1992). J. Cryst. Growth, 122, 161-167.
Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com 250 Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com
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Methods, Procedures, Technical continued...
Skin on Drop
Are you seeing skin form on the surface of your drops containing sample and reagent and wondering what to do?
Skins on the drop are considered bad. These skins are believed to be a layer of denatured protein. Skins can slow down the rate of vapor diffusion.
Pick away the skin before mounting your crystal. If the skin wraps around the crystal, it can affect the limit of diffraction.
To avoid skins, one can try crystallize under oil.
1
Either put a few l of oil on top of the sitting drop, or use the microbatch under oil method.
Keep in mind that covering the sitting drop with oil will affect the rate of vapor diffusion equilibration. The following oils are ranked in order of the rate of
equilibration allowed by the oil: Paraffn Oil (slowest rate), Als' Oil (medium rate), Silicon Oil (highest rate).
Reference:
1. Pearl, B.OHara, R.Drew, S.Wilson. Crystal Structure of AmiC: the controller of transcription antitermination in the amidase operon of Pseudomonas earuginosa. EMBO Journal 13. (1994), pp 5810-5817.
Vibration
Should one be concerned about vibration during crystallization?
1. "Many years ago at Imperial College we did a fairly systematic experiment on vibration using lysozyme (hanging drops in Linbro trays). We placed one
tray in the basement on a very solid concrete base, one tray was placed in an incubator, frequently used and known to vibrate a little (with frequent door
opening and closing), and the third tray was placed on a rather strongly vibrating metal plate hanging from a ceiling on a piece of wire. The vibrations were
produced by a small motor with an acentric piece of metal attached to its rotor. The trays were examined at regular intervals. Crystals grew in all of them.
The basement tray gave fewer but bigger crystals, but this may have been a small temperature effect - temperature was not controlled very well. Crystals
from all trays were nice, they diffracted to the same resolution, with virtually identical Wilson distribution. After this experiment we stopped worrying about
vibration." Tadeusz J. Skarzynski, 2006 CCP4 Bulletin Board
2. "...This is not to say, however, that absolute silence or stillness must prevail in the presence of growing crystals. At the Massachusetts Institute of Technol-
ogy all the tRNA and protein crystals were grow in a cold room that had a giant compressor attached and contained, as well, several ancient centrifuges
and shaker baths. One could observe standing waves in the reservoirs of all the vapor diffusion chambers and frequently could scarcely converse above the
sound. Whether this had a positive or negative effect, we could not be sure. It is more likely that dramatic changes in the environment, as those caused by
handling, are more disruptive than ambient conditions themselves." Alexander McPherson (1976) "The Growth and Preliminary Investigation..." in Methods
of Biochemical Analysis, Vol. 23, page 284.
3. CCD Video Observation of Microgravity Crystallization of Lysozyme and Correlation with Accelerometer Data. E. H. Snell, T. J. Boggon, J. R. Helliwell, M. E.
Moskowitz and A. NadarajahActa Cryst. (1997). D53, 747-755.
Synopsis: Stepped growth rates and crystal movements have been observed by CCD video in microgravity lysozyme protein crystallization experiments
which correlate with `g-jitter' accelerations, especially of low frequencies. The spurts and lulls of crystal growth may, therefore, explain the residual mosaic
block structure seen in protein crystal mosaicity and topography measurements.
Additional Comment by James Holton: They saw a very clear conneciton between vibrations and crystal growth rates and indeed crystal quality. Things like
"astronaut exercising" coincided with increased crystal growth rate a short time later. So, it would appear that even in the most controlled environments
vibration control can be a challenge, but at least on the Shuttle, where everything gets logged, you can (could) look for relationships.
And a comment from Eddie Snell: Basically, for vibration there are two factors, frequency and magnitude. Any high magnitude vibration is bad, i.e. the sud-
den short duration, hopefully low frequency one when you drop your tray :) For the same magnitude, low frequencies (a few Hz) are worse than higher.
frequencies (mains frequency). High frequencies are quickly damped in a liquid environment. Measuring vibrations is fairly easy, translating that measure-
ment into something useful is not. In the case James mentioned, we were looking at the reduced acceleration environment. Certain crystallization processes
lasted over a longer time period than on the ground - in this case the presence of a halo of depleted solution around the crystal caused by the diffusion
limited transport of the protein to the crystal face. Every time an astronaut exercised, this halo broke down and a growth spurt occurred (except for one
astronaut who didn't exercise very hard at all, however he/she shall remain nameless). On the ground, this depletion zone is only there for a short period of
time if at all. Transport and therefore growth is dominated by convection - the density differences cause fow of sample across the growing crystal face. Any
effect from vibration at high frequency in a liquid medium would only be meaningful if the magnitude of that vibration was large. Diffusion processes and
surface tension effects in the drop and Brownian motion and surface dynamics on the crystal face probably dominate the crystallization process at the mac-
romolecule scale. Although I don't think vibration will be too big of an issue unless the magnitude is large I personally like small bench top Peltier controlled
continued next page...
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Methods, Procedures, Technical continued...
Vibration continued...
forced air incubators. I can rule vibration out as a variable. Vibration isolation may make a difference with large magnitude vibrations, however vibration
reduction would be cheaper. The best thing to do is try a simple experiment with a few of your favorite proteins. Set up a tray in one of the rooms if a col-
league has one available and duplicate trays in other incubators set at the same temperature. If there is any big difference you have your answer (assuming
it is not light, temperature control or a million other differences). Frequently I have often found that nice physical explanations in crystal growth are often
poor approximations and later prove to be incorrect - a little shaking might be good after all. Perhaps that is why there are so many crystallography groups in
California :)
His Tag and Ni Help Phasing
Structure of Mycobacterium tuberculosis mtFabD, a malonyl-CoA:acyl carrier protein transacylase (MCAT). Hemza Ghadbane, Alistair K.Brown, Laurent Kre-
mer, Gurdyal S. Besraa and Klaus Futterera. Acta Cryst. (2007). F63, 831835. The crystal structure of M. tuberculosis mtFabD, the mycobacterial MCAT, has
been determined to 3.0 A resolution by multi-wavelength anomalous dispersion. Phasing was facilitated by Ni2+ ions bound to the 20-residue N-terminal
affnity tag, which packed between the two independent copies of mtFabD.
Crystallization of (insoluble) Ligand Protein Complexes
Crystallizing ligands with limited or no water solubility can be tricky. Here are some suggestions.
A good review on the crystallization of protein ligand complexes can be found in Crystallization of proteinligand complexes, Annie Hassel et al, Acta Cryst.
(2007). D63, 7279.
Solubilize the ligand in DMSO so it is maximally concentrated (100mM works fne). Add enough ligand to achieve two to three fold excess ligand to protein.
Keep the DMSO concentration to no more than 3% to avoid damaging the protein. Set crystallization experiment using this sample. If you cannot achieve a
high enough stock concentration of DMSO to be below the 3% threshold, dilute you protein in the storage buffer to approximately 1mg/ml. Add compound
to two to three fold excess, incubate and co-concentrate to the desired concentration. This may help to avoid the DMSO shock. Alternatively one can incu-
bate the concentrated protein with the compound solubilized in water for 24 to 48 hours and solubility of the ligand will be suffcient to complex with the
protein. (Carsten Schubert ccp4bb December 2007)
One method that worked for me was to dissolve my ligand in 100% DMSO, as suggested in the previous response, then add a 3 molar excess of ligand to
protein so that the fnal concentration of DMSO in the protein-ligand solution was no greater than 10% - of course the maximum concentration of DMSO
that your protein can suffer will be protein specifc but you could investigate this by incrementally adding DMSO to just a solution of your protein at your
working crystallization concentration then measuring scattering at 600nm in a spectrophotometer to determine the critical DMSO concentration that causes
your protein to precipitate (if you have suffcient protein to 'waste'!). If your ligand binds tightly to your protein at an equimolar concentration you can then
remove excess ligand and DMSO by passing your sample through a G25 Sephadex column. (Rob Hussey ccp4bb December 2007)
If you grow crystals in polyethylene glycols or similar reagent you might try to solubilize the compound in a small amount of this reagent. This is helpful if
you want just a 1:4 protein:ligand ratio. Sometimes solubility is low even in 5% DMSO (or diluted solutions of glycerol, alcohols and similar molecules). In
these cases setting up drops in the presence a saturated solution and some precipitate of the compound may also lead to good co-crystals. As one molecule
passes from the saturated solution to the bound state, a new molecule is solubilized from the precipitate, which gradually dissolves and passes from the
solution to its binding site in the protein. Indeed, this sounds like a soaking experiment and it works well if the compound is colored, so that you can see
if the crystals actually become of the same color. Just remember to wash them thoroughly before measurements, in order to remove traces of the ligand
precipitate that would result in poor diffraction. (Marco Mazzorana ccp4bb December 2007)
We routinely obtain structures from protein solutions with a big pellet of ligand in the bottom of the tube. For co-crystallizations we add 1mM compound to
a 0.3mM solution of the protein and incubate overnight. Many of the compounds are only soluble to 50 micromolar, so we get a lot of precipitate. The next
day, we spin the tube at high speed, and use the supernatant for crystallization trials. We have started from 100 mM stocks in 100% DMSO or ethanol. This
has worked for compounds ranging for picomolar to micromolar affnity, which surprised us, but it worked. (Kendall Nettles ccp4bb December 2007)
Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com 252 Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com
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Methods, Procedures, Technical continued...
Glutaraldehyde to Differentiate Salt from Protein Crystals
Place the crystals in a low ionic strength buffered solution containing 0.2-.2.0% v/v Glutaraldehyde. Protein crystals will quickly be fxed (faster than they
dissolve) into a light golden, gelatinous lump. Sometimes the crystals retain a crystal-like shape, other times the crystals leave just a rubbery blob. In contrast
to protein crystals, salt crystals should dissolve over time and should not be colored.
One can add a small amount of 2% v/v Glutaraldehyde to the protein drop. Protein crystals in the presence of Glutaraldehyde will then turn a light golden
color. Crystals fxed like this can then be put into a low ionic strength solution where salt crystals should dissolve. You can easily transfer Glutaraldehyde
into a protein drop by vapor diffusion by adding it to the reservoir to make it 2-3% v/v.
A caveat to using this method is that there should be no free amines around other than on the protein (i.e., no Ethanolamine or Tris buffer, no Ammonium
ions, etc.).
Note: Glutaraldehyde is toxic, causing severe eye, nose, throat and lung irritation, along with headaches, drowsiness and dizziness. It is quite volatile with a
strong, obnoxious odor.
A suggestion from Michael Garavito, Michigan State University.
Making a Bigger Crystal or Changing Crystal Form
1. Screen additives. See Hampton Research Additive Screen

and Detergent Screen

.
2. Use a crystallization screen as an additive screen. Make the reservoir 90% original reagent plus 10% crystallization screen reagent
(Crystal Screen

, Index

, etc). Repeat using the entire screen.

3. Change the crystallization method: Sitting drop, hanging drop, microbatch, dialysis, free interface diffusion, etc.
4. Try limited proteolysis. Reference: In situ proteolysis for protein crystallization and structure determination. Nature Methods - 4, 1019 - 1021 (2007).
5. Add fresh protein to the drop once crystal growth has ceased.
6. Add water to the drop once crystal growth has ceased. Crystals may dissolve and grow larger or a new crystal may form.
7. Try the Silver Bullets

screen from Hampton Research. Reference: Searching for silver bullets: An alternative strategy for crystallizing macromolecules.
Alexander McPherson and Bob Cudney. Journal of Structural Biology 156 (2006) 387-406.
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Optimization
Sialidase to the Rescue
Sialidase I from Clostridium perfringens (Glyko, Inc. Rosedale, NY, USA) was used by Ogawa et al (Acta Cryst. (2003). D59, 1831-1833) to remove terminal
sialic acid residues and minimize the heterogeneity of the glycosyl structure on the protein and enhance crystallization. The protocol utilized 1 mU of Siali-
dase I in 50 microliters of 50 mM Tris-HCl buffer pH 6.8 at 310 K for 12 hours.
Substituting Sodium Malonate for Ammonium Sulfate
Xing and Xu (Acta Cryst. (2003) D59, 1816, 1818) recently describe the crystallization of the PX domain of cytokine independent survival kinase in Am-
monium sulfate. Optimization of the crystallization seemingly involved the subtle manipulation of several crystallization variables, including using drop ratios
(1:2 drop ratio), the removal of DTT to allow crystal growth after nucleation, as well as the substitution of Sodium malonate for Ammonium sulfate. The
authors reported the Sodium malonate dramatically increased the reproducibility of the crystals and also acted as a good cryoprotectant. Crystals grown
from 2.0 M Sodium malonate could be cryoprotected directly from the drop. The crystals grown in Sodium malonate were more resistant to physical shock
compared to those grown in Ammonium sulfate.
Sample Purication as an Optimization Variable
If one is able to obtain crystals of a protein but the crystals do not diffract well, try further purifcation of the sample and repeat the crystallization condi-
tions. Oftentimes crystals do not diffract well because of impurities or aggregates present in the sample. Impurities in the sample can cause dislocations and
defects in the crystal, which can lead to poor diffraction. Further purifcation can sometimes remove these impurities and aggregates, resulting in crystals
with improved diffraction. Oftentimes the reason crystals do not diffract can be traced to the molecules in the crystal not being well ordered in the lattice.
Therefore, there is conformational fexibility or the molecules do not take a desirable or fxed conformation because the lattice forces are too weak. Or
sometimes there are severe defects in the crystal lattice, which lead to disorientation.
Drop Size
Try increasing the drop size to grow larger crystals.
Remove Aggregates
Centrifuge the sample at 2,000 g for 10 to 15 minutes immediately prior to setup to remove aggregates and amorphous material.
Using Additives
During optimization of crystallization conditions, examine any additive that might for some reason tend to stabilize or engender conformity by specifc
interaction with the macromolecule.
Saving Kits
To reproduce crystallization conditions from a screen without taking additional solution from the screening kit (and making all your lab partners want to do
evil things to you), simply set an additional drop or drops on the cover slide next to the original drop. One can set multiple drops on the same cover slide
over the same reservoir. Pipet the sample onto the cover slide, then pipet the precipitant from the reservoir into the sample drop. Lastly, seal the cover slide
over the reservoir.

Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com 254 Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com
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Optimization continued...
Varying Ionic Strength
It has been suggested that lattice contacts within a crystal can be manipulated using ionic strength, which in turn can be used to infuence crystal morphol-
ogy. Rather than the type of salt used (Sodium chloride, Sodium acetate, Sodium citrate, etc.), one may manipulate the concentration of the salt or change
crystal morphology. Typical salt concentrations to be screened for such manipulation are 0 to 500 mM.
Explore All Angles
Explore as many optimization opportunities for crystallization as possible. Optimize every hit from a screen. When you reach an optimization dead end for a
particular hit, review the screens again and optimize crystal leads from other hits. Optimize different hits simultaneously if sample availability permits. This
can save time if one hit leads to a dead end (poorly diffracting crystal, non-isomorphous derivative, etc.).
Beauty is Only Skin Deep
One's tendency is to optimize the largest, best looking crystals. Common sense. However, there are plenty of reports where small, miserable, ugly crystals
have eventually produced the best data, so do not ignore the ugly ducklings.
Control
Control is a good thing. Having a handle on the variables which affect the crystallization and the ability to control these variables is key to the reproducible
growth of high quality crystals.
Fresh Ligands
When working with ADP, ATP, NAD, NADH or other chemically unstable ligands as crystallization additives, be sure to use freshly prepared working solu-
tions or properly stored stocks. These agents are sensitive to degradation over time which can infuence the outcome of a crystallization experiment. Also,
consider the pH of these ligands in solution since the pH of the additive agent can infuence the fnal pH of the drop. It might be necessary to titrate the
ligand solution before addition to the drop.
Increasing Crystal Size
How might one increase the size of a crystal which is grown from dialyzing the protein into pure deionized water? The following replies were posted on the
cc04bb (http://www.ccp4.ac.uk) on October 21, 2005.
George Kontopidis: If I was you the frst thing I will try is to increase protein concentration. Also after crystal formed you can take the solution to room
temperature that also may help.With the existing crystals just open the container or the vial where the crystals formed to allow further evaporation ( do not
let it dry out) which will result to increase of protein concentration and hopefully increase crystal size.
A.ErtugrulCansizoglu: You might try vapor diffusion against water. As such you keep you protein sample in a mild-salty condition- where it doesn't actually
crystallize, then set up your drops using pure water. This worked for a case in our lab. Slowing down the crystallization made the crystals bigger.
Sameer Velankar: I once crystallized a protein in similar conditions. For that matter many Thymidylate synthase crystallize by decreasing the salt concentra-
tion. You can add very little salt (15-25mM) in the drop and keep water or 2-5mM salt in the well. This controls the growth of the crystals and if you are lucky
you will get bigger crystals.
Christopher F. Snook:I would suggest increasing the drop volume, i.e. going from say 4 to 8 microliters. This has a two-fold effect: an increase in the protein
available for crystallisation and a reduction in the equilibration of the drop with the well solution. Another possiblity would be to increase the concentration
which may increase the nucletaion at the expense of crystal growth.
deacona@mail.rockefeller.edu: I have never encontered such a situation, but maybe some microdialysis from your storage buffer against pure water may
help somewhat? I believe this is a common strategy for antibody crystallization. As usual, micro/macro-seeding, varying protein concentration etc may also
be of use. Just some thoughts...P.S. These may be of some interest J Biol Chem. 1970 May 25;245(10):2763-4 A crystallographic investigation of a human IgG
immunoglobulin.Edmundson AB, Wood MK, Schiffer M, Hardman KD, Ainsworth CF, Ely KR.
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P A G E 2 1 - 2 9
Optimization continued...
Increasing Crystal Size...continued
J Mol Biol. 1997 Dec 19;274(5):748-56. Use of organic cosmotropic solutes to crystallize fexible proteins: application to T7 RNA polymerase and its complex
with the inhibitor T7 lysozyme. Jeruzalmi D, Steitz TA.
Oleg Tsodikov: I would still spin down the crystals (or dissolve them in the presence of small amount of salt or glycerol in and some buffer) and then screen
in a variety of different conditions.
David Aragao: We had, some years ago, a very similar case (salting inn?). Our crystals started to appear in the purifcation process everytime the ionic
strenght decreased. The fnal crystals were big and we got 1.2 resolution from them. I know there are nice comercial dialysis buttons nowadays but we
made our ones at the time. Maybe the description of our optimization process can help you (see materials and methods). Acta Crystallogr D Biol Crystallogr.
2003 Apr;59(Pt 4):644-53.
jjwarren@duke.edu: I would recommend that you check out Jeruzalmi, D. and Steitz, T.A. (1997) Use of organic cosmotropic solutes to crystallize fexible
proteins: application to T7 RNA polymerase and its complex with the inhibitor T7 lysozyme. J. Mol. Biol., 274, 748-56. Their methods are generally appli-
cable, but they focus heavily on proteins that crystallize at low ionic strength...
Christopher Colbert: Seriously, you might want to consider screening for additives.
Ewa Skrzypczak-Jankun: Try salt - increasing the ionic strength often makes crystals more bulky. Also adding salt will change saturation point and can slow
crystallization If nothing else NaCl is CHEAP. Do you have any information about affnity to ions? what your protein likes and dislikes? digging in the litera-
ture can provide valuable clues.
Stephen Prince: You could try slowing the rate of cooling - there was an old recipe for insulin crystallization that involved slow cooling. One used a thermos
fask containing water at above room temp, the protein mix was foated in this in a small test tube, the thermos was sealed and placed in a polystyrene box
and left undisturbed for a week or so, crystals grew in the tubes as the solution cooled. You could try the same trick in a cold room ...
Jonathan Caruthers: I've seen the same effect on at least two occasions - actually I believe hemoglobin was frst crystallized by dialyzing blood in water if
memory serves. In any event, we managed to grow diffractable crystals simply by removing them from the fridge, partially melting them, and putting them
back in the fridge to re-grow. This works pretty well.
William Scott: The main concern is stability. If you can fnd a more conventional crystallization, it might ultimately make your life easier.
The good thing is you know it will crystallize readily. (I assume this is a good thing to know, unless you are trying to do NMR)
Raji Edayathumangalam: If you want to pursue these very crystals - try oil, which retards nucleation and you get bigger (and often better) crystals. We've
had a case before where crystals falling out of the protein solution weren't the best bet. They were inherently disordered - not the optimal crystals. Crystal
screens yielded crystals in other conditions that diffracted to higher than 2Ang.
Jon Caruthers : actually, to follow up here, I grew crystals of a MBP-protein fusion construct using diet-coke as a buffer. It turned out that these crystals
diffracted better than anything grown with any other buffer of similar pH (2 vs. ~3.5 with other buffers), so maybe there's something to this idea of using
CO2 as a buffer.
Tassos Papageorgiou: Try dissolving the crystals in salt and dialyse them afterwards against water might help. Have you try to see whethre crystals grow in
low ionic strength?
David Waterman: I had a similar case with a protein which crystallised on a drop in ionic strength. The protein was soluble in ~100 mM K Phos, and started
forming crystals when this concentration was reduced by diluting with water (or glycerol, which had the bonus of being a cryoprotectant). The biggest crys-
tals (blocks with sides up to 0.3-0.4 mm ) were obtained after setting up a hanging drop of concentrated protein over a reservoir of water alone (no mixing!)
and allowing this drop to grow slowly by vapour diffusion. These crystals were diffcult to cryo-protect though, so a compromise including glycerol in the
reservoir and mixing with the protein drop was found. It was certainly much easier than fddling with dialysis buttons, though I might have been lucky.
Peter Moody: I crystallised a DNA repair protein by dialysis to remove the salt that kept it soluble. Crystal quality was much improved with 300mM sugar
(Maltose seemed the best).
Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com 256 Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com
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Optimization continued...
No Diffraction - What to Try?
a. The protein may have a high degree of structural fexibility and therefore structural heterogeneity. Try screening ligands, additives, co-factors and other
small and large molecules that might help the protein assume a more rigid conformation.
b. Structural variations in the protein due to enzymatic modifcation, partial deamination, partial oxidation, other post translational modifcations can lead to
sample heterogeneity. Analyze your sample and conditions from expression to solubilization to purifcation to crystallization to identify what might lead to
sample heterogeneity. Remove it or change it. If working with an enzyme, bind an inhibitor or ligand to the enzyme in an attempt to lock into a more rigid
and homogeneous conformation. Reduce the number of steps and the amount of time between expression and crystallization.
c. If the crystal is growing overnight, the high rate of crystal growth may indicate a high level in the number of defects in the crystal lattice. Slow the rate of
crystal growth. Reduce reagent concentration or protein concentration. Layer silicon oil over the reagent reservoir to slow vapor diffusion. Use less reagent
in the reservoir. Evaluate drop dilution and drop ratios. For example, dilute the protein 1:1 with your sample buffer. Set vapor diffusion experiments with
diluted sample:reagent drop ratios of 1:1, 2:1 and 3:1. The fnal protein concentration will remain similar between the original and diluted samples but the
path of equilibration will be different and the rate will be lower than for the original drop.
d. Mount the crystal, pre cryo, in a capillary and test for diffraction to confrm that cryo is not causing the poor diffraction. If room temperature data is better
than cryo, your cryo method and cryo procedure might beneft from further refnement.
e. Screen additives.
f. Screen crystallization kits as additives. Mix 70 to 85 l of the optimized crystallization reagent with 30 to 15 l of solutions from your favorite Hampton
Research screen (Crystal Screen

, Crystal Screen 2

, Index

, SaltRx

, etc.) to create a 100 l solution. Mix thoroughly. Pipet 100 l of this solution into a
96 well sitting drop plate. Create a drop mixing equal amounts of reagent and sample, seal and allow to equilibrate. Varying the ratio of optimized reagent:
screen reagent and drop ratio can also be tried. Watch out for false positive salt crystals as these random mixes may create insoluble salts. Look in the
reservoir for salt precipitates or crystals. If one prefers to use 24 well plates, simply change the ratio of optimized crystallization reagent:screen reagent. For
example, in a Cryschem

Plate, try 350-450:150-50. In a VDX

Plate, try 750-900:250:100.

References:
1. Personal communication with Annie Hassell at GlaxoSmithKline, circa 1996.
2. Acta Cryst. (2005). D61, 646650. Crystallization of foot-and-mouth disease virus 3C protease: surface mutagenesis and a novel crystal-optimization strategy. James R. Birtley and Stephen Curry.
Desiccating Crystals to Improve Diffraction
Desiccating, or drying a protein crystal can be a way to salvage or improve the diffraction quality. The procedure consists of removing a non-diffracting
crystal from the X-ray beam, plunging it into a soaking solution made of the original crystallization reagent supplemented with a suitable cryoprotectant
(glycerol, ethylene glycol, MPD, PEG 400, etc.) and then allowing the drop to dry in the evaporating sitting or hanging drop for 15 minutes to several hours.
Try 1 ul cryo reagent plus 9 ul of original crystallization reagent for the drop. Remount the treated crystals and examine for diffraction. The procedure is
originally described by Chantal Abergel. Reference: Spectacular improvement of X-ray diffraction through fast desiccation of protein crystals. Chantal Aber-
gel. Acta Cryst. (2004). D60, 1413-1416.
Dissolve and Restore Protocol for Growing Larger Crystals
Impurities adsorbing onto the crystalline surface can prevent further growth of a crystal. Repair and growth of an impurity poisoned crystal is possible by us-
ing a dissolve and restore protocol described by Plomp et al (2003). By temporarily applying undersaturation conditions, an impurity adsorbed layer can be
removed, which followed by saturation conditions can revive crystal growth with the potential for crystal growwth without the incorporation of impurities.
Regular cycles of short periods of undersaturation followed by longer periods of supersaturation can be applied to dissolve and restore, with the potential
of a larger crystal with fewer impurities and defects. Temperature manipulation (cycling) for samples with temeprature dependent solubility is a convenient
approach. Altering and cycling sample and reagent concentration is another approach (Heinreichs et al (1992), Koeppe et al (1975), Przybylska (1989)).
References:
1. Plomp et al (2003). Repair of impurity-poisoned protein crystal surfaces. Proteins: Structure, Function, and Genetics 50:486-495.
2. Heinreichs et al (1992). Growth of single protein crystalsin a periodically solubility gradient: description of the method and rst results. J. Cryst. Growth 122:186-193.
3. Koeppe et al (1975). A pulsed diffusion technique for the growth of protein crystals for X-ray diffraction. J. Mol. Biol. 98: 155-160.
4. Przybylska (1989). A double cell for controlling nucleation and growth of protein crystals. J. Appl. Cryst. 1989:22:115-118.
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P A G E 2 3 - 2 9
Optimization continued...
Heavy Atoms and Salt Reagents
Heavy atom derivatives can sometimes be diffcult to obtain in the presence of sulfate salt crystallization reagents such as ammonium sulfate or lithium
sulfate.
However, diffcult does not mean impossible, so be sure to screen heavy atoms in the salt based condition before trying to cure a problem that might not
exist.
When one is unable to obtain succcessful derivatives in the presence of sulfate salt based crystallization reagents, one might try transferring the crystals to
high concentrations of a similar acetate salt based crystallization. For example, Ammonium acetate instead of Ammonium sulfate, or Lithium acetate instead
of Lithium sulfate. This will maintain the high ionic strength of the crystallization reagent yet increase the solubility of some heavy atoms. Acetate, in higher
concentrations can also act as a cryo salt.
Try replacing the original salt with Sodium malonate or Tacsimate. Malonate: a versatile cryoprotectant and stabilizing solution for salt-grown macromolecu-
lar crystals. T. Holyoak, T. D. Fenn, M. A. Wilson, A. G. Moulin, D. Ringe and G. A. Petsko. Acta Cryst. (2003). D59, 2356-2358.
Try halide soaking using KI or KBr. Novel approach to phasing proteins: derivatization by short cryo-soaking with halides Z. Dauter, M. Dauter and K. R.
Rajashankar Acta Cryst. (2000). D56, 232-237.
One might also consider trying 100 mM Cobalt hexammine or Iridium hexammine as derivatives in the presence of acetate salts.
Try cross-linking. A gentle vapor-diffusion technique for cross-linking of protein crystals for cryocrystallography. Carol J. Lusty. J. Appl. Cryst. (1999).
32, 106-112.
Sulfate Ions in the Active Site
Problem: Enzyme crystals grown in Ammonium sulfate having trouble binding ligand.
Solution: This is not an unusual situation with sulfate, and, yes, sulfate often occupies phosphate binding sites, particularly when you're work with 1-3 molar
concentration of Ammonium sulfate. The simplest way to remove sulfate from the crystal is to transfer to high concentrations of citrate. From my own
graduate work on GPDH in the 1970's, we just transferred the crystals from 3.0 M AS to 1.44 M Sodium citrate. While the crystals weren't stable in citrate for
long periods of time (days), that is not an issue today with cryocrystallography and fash-freezing. You might even try formate or malonate, which can have
a modest cyroprotectant behavior. Another, and perhaps more relevant, method is the one devised by Bill Ray in the late 1980's for phosphoglucomutase
(PGM). Apo-PGM was crystallized in high AS, which caused the same problems as you are experiencing. Ever the perfectionist, Bill devised a systematic
method to transfer PGM crystals from AS to a PEG solution that allowed the formation of enzyme complexes (Biochemistry 30, 6866, 1991). Bill also looked
at glycine as a replacement as well. So there are a number of ways to "desalt" crystals without resorting to cross-linking, while preserving good diffraction
characteristics. Michael Garavito, Michigan State University.
Vary the Method of Crystallization
Using the crystallization reagent that produced the initial hit, or reagent formulations which have been partially optimized, set additional crystallization
experiments using other methods and techniques. If sitting drop vapor diffusion was used to produce the initial hit, try hanging drop, vapor diffusion,
microbatch, free interface diffusion, or dialysis. Try different plates and different apparatus using the same method to discover whether the difference in
equilibration will produce a better crystal, a crystal in a shorter period of time, or a different crystal form. The volume of the chamber or well, the space
between the drop and the reagent, the shape of the chamber and the well, and also the method and time required to set the experiment may affect the
outcome of the crystallization experiment.
The path is as important as the starting and ending points in determining ideal nucleation and growth conditions.
Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com 258 Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com
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Optimization continued...
Omit the Irrelevant
If the crystallization reagent producing a hit is a complex mixture containing, for example, salt, detergent, additive, metal ions, ligands, reducing agent,
EDTA, cryogen, or other chemicals, prepare optimization reagents based on the hit, but systematically omit each of the components, one at a time. Which
chemicals appear to make a difference in the crystal size, number and quality? Omit from future optimization experiments any chemical which appears
to be irrelevant. Identify and remove irrelevant chemicals before adding the evaluating new chemicals. As with the sample buffer, keep the crystallization
reagent as simple as possible. Include only chemicals which promote and enhance sample stability, homogeneity and crystallization.
Optimization is an Art
The skill of crystal optimization is an art form that improves with effort, thought, discussion, refection, reading, learning, trial and error, patience, tinker-
ing and time.
The path of optimization is rarely linear and the variables affecting crystallization are not independent. This makes it impossible to sample all crystalliza-
tion variables and evaluate them at fne intervals.
Optimization may follow a main trail, but there are typically many branches in the trail to consider, explore and evaluate. And typically, time and sample
availability do not allow one to explore each and every branch.
So one is left with the question of what they should try or what they should do next to increase the size or improve the quality of of the crystal. And there
are usually plenty of recommendations to consider. But which recommendations should be considered and tried and in what order or priority, and which
ones should be ignored for now and tried later, or perhaps discarded altogether?
It is the art of an experienced crystal grower to consider the results of systematic experiments of many different samples over much time and use this
knowledge base to synthesize the best plan for growing perfect crystals.
Optimization can be tedious and require numerous iterations before one uncovers which combination chemical and physical variables produce the
perfect crystal.
Optimization is an art and a technical skill. As such, it is dynamic and improves in direct proportion to the number of challenges, the degree of the chal-
lenges, and experience.
A good crystal grower never is, they simply get better and better and better...
Build a Tool Box
A skilled craftsman knows they cannot do their job without the proper tools. Maybe some jobs can be accomplished with one or two common and simple
tools, but a skilled craftsman, over time, will build not only experience, but also a tool box. A tool box of both general and specialized tools that will allow
them to handle all tasks in an effcient manner. And over time, a skilled craftsman gains appreciation of the importance that quality, reliability and support
bring when deciding which tools to acquire.
A crystal grower also needs to build a tool box. This tool box will consist of apparatus and devices, plates and seals, kits and reagents; some general, some
specialized. Initially the crystal grower will want to acquire tools that have general utility. Later, more specialized tools can be added so that when the need
arises, these tools will be available to use and evaluate. Access to a useful and broad portfolio of tools ensures the crystal grower will have the ability to
evaluate those variables which could lead to the perfect crystal.
You can only use and evaluate what you have and know how to use.
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P A G E 2 5 - 2 9
Sample Preparation
Imidazole and Crystallization
Should one remove 1.0 M Imidazole from the sample after elution from a nickel column? In many cases it is good to have (an) additional purifcation step(s)
following the elution of the sample from a Ni column. Size exclusion chromatography (SEC) is a convenient and effective way to remove the Imidazole. If
one fnds that Imidazole is an important variable in maintaining sample homogeneity, one can also simply reduce the concentration from 1.0 M to a more
reasonable level (0.02 0.2 M) for inclusion of crystallization reagents. In some cases, a protein might not crystallize in the presence of Imidazole. To
remove the Imidazole, one can try precipitating the protein in Ammonium sulfate and resuspending the sample in low salt buffer and purify by ion exchange
or SEC. One can also remove Imidazole by dialysis. In one instance (personal communication from Artem G. Evdokimov) decent crystals could only be
obtained using 0.6 M Imidazole/acetate buffer and two Imidazole molecules were seen occupying hydrophobic spots on the protein surface. Sometimes
the reason Ni column purifed proteins precipitate without Imidazole is that the Ni ions leak from the Ni column. The presence of Imidazole may have been
chelating this excess and trace Ni and once removed, the remaining Ni may cause the protein with a His tag to aggregate in solution. One may be able to
remove this excess Ni using a Ni chelating resin (Hampton Research catalog number HR2-312). Or one can leave the Imidazole buffer with the sample in
a more reasonable concentration (0.02 0.1 M) or try Citrate buffer (chelator) or EDTA. If you are working with a metalloprotein and need metals in the
sample, using the chelating resin rather than leaving a chelating reagent in the sample is obviously a better choice.
Heat Treatment
If a protein does not crystallize or gives poorly diffracting crystals, one might try heat treatment of the sample. This is a quick experiment with minimal
equipment required. The procedure can reduce and/or eliminate protein that is not folded properly from the sample. Place the sample at 4C for one hour.
Incubate the sample at 37C for one hour. Centrifuge the sample or flter using a 0.22 micron flter. Screen the heat treated sample for crystallization. The
time course and temperature may require optimization for best results. One can monitor the heat treatment using dynamic light scattering or an activity
assay. Tip from Annie Hassell, GSK IUCr 2005.
Sample Too Soluble To Crystallize
The "solution" we hit upon for a problem protein with a high solubility profle was to cocrystallize it with an Fab fragment. Not the easiest route, but it
worked. Catherine L. Lawson, Rutgers University. Reference: Li H, Lawson CL. (1995) Crystallization and preliminary X-ray analysis of Borrelia burgdorferi
outer surface protein A (OspA) complexed with a murine monoclonal antibody Fab fragment. J Struct Biol. (115 )335-337.
Dealing with Glycosylation
When having trouble removing glycosylation from a protein before crystallization, try a sequential exo-glycosidase(s) then endo-glycosidase treatment in
order give the endoglycosidase better access. Or, try to prevent glycosylation in the frst place and try glycosilation inhibitors (tunicamycine for example
inhibits N-glycosilation). Another option is to try the expression of the protein in GnTI(i) HEK293S cells which are unable to synthesize complex N-glycans
(Proc Natl Acad Sci U S A. 2002 Oct 15;99(21):13419-24. Reeves PJ, Callewaert N, Contreras R, Khorana HG. Structure and function in rhodopsin: high-level
expression of rhodopsin with restricted and homogeneous N-glycosylation by a tetracycline-inducible N-acetylglucosaminyltransferase I-negative HEK293S
stable mammalian cell line). One more alternative suggestion is to use your original expression system, but with Asn-Asp mutations of the glycosylation sites;
assuming of course the mutant proteins will still be soluble which is straightforward to test. If the protein is resistant to treatment with Endo H or PNGase F
under native conditions, try the Endo H treatment under only slightly denaturating conditions. Try screening a gradient of denaturating conditions to see if
you can fnd a concentration that allows deglycosylation while not unfolding your protein.
There are reports where a fully glycosylated protein is able to be crystallized. So it may be worth a try to crystallize the glycosylated form. If the sample will
still not crystallize, try deglycosylation, preventing glycosylation, or another cell line.
Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com 260 Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com
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Sample Preparation continued...
Deglycosylation Summary
Deglyosylation Summary
(built on tips from the CCP4 community)
Glycoproteins are proteins to which a carbohydrate chain is covalently attached. Proteins modifed in this way in many cases present a considerable
challenge to X-ray structure analysis, because they do not easily form crystals. This is because the attached carbohydrate chains are often heterogeneous
and fexible and interfere with the formation of crystal contacts. A well-known strategy to tackle this problem is to deglycosylate the glycoproteins prior to
crystallization (see below for practical tips). This is usually done with endoglycosidases such as PNGase F (resulting in virtually complete removal of all N-
linked glycans) or Endo H, Endo F1, F2, F3 (for partial deglycosylation). Also exoglycosidases can be used, alone or in combination with endoglycosidases.
Alternatively, the glycosylation sites can be mutated (substitution of S/T to Ala or N to Gln or Asp in the conserved NXS/T motif). Expression in lower
organisms like E. coli or Pichia pastoris is also an option, or - if a less radical procedure is required - genetically modifed expression systems can be used
that do not contain certain glycosyltransferases (e.g. CHO Lec cells).
Practical tips for enzymatic deglycosylation:
Several companies like Sigma-Aldrich and ProZyme have excellent theoretical and practical information about deglycosylation on their web pages, which
are well worth studying. Both N- and O-glycosylation occur naturally in proteins, with N-glycosylation being the more common modifcation. N-glycans can
generally be removed rather effectively by a single enzyme, PNGase F, whereas this is not the case for O-glycosylation, where several enzymes have to act
in concert to exert the same effect. Deglycosylation can be tried both under native and denaturing conditions. While native conditions are in general pref-
erable for crystallization purposes, deglycosylation under denaturing conditions is more effective and should for this reason always been done in parallel as
a positive control. To achieve higher effciency even under native conditions, one may experiment with using higher temperatures (up to 37C, instead of
0-4C) and/or longer reaction times (1-5 days; preferably adding some Sodium azide to the mixture to prevent bacterial growth).
In some cases, native deglycosylation does not work well. In these cases, one may try to deglycosylate the protein under denaturing conditions and then
refold the protein. Alternatively, one may use only slightly denaturing conditions, by applying various mild detergents (like b-octyl-glucoside, Chaps, Triton
X-100, SDS, etc.). One can also try to incubate the reaction mixture in a sonicating water bath. Yet another option is to deglycosylate the protein only
partially using exoglycosidases (neuraminidase, galactosidase, etc.) instead of endoglycosidases.
In practice, one often starts out with a protein solution (concentrated to 0.5 to 5 mg/ml in water or a suitable buffer) and adds the glycosidase of choice
(e.g. PNGase F, Endo H, Endo F1, F2, F3, neuraminidase or enzyme kits) in ratios varying from ca. 1:15 to 1:2000 (w:w). The deglycosylation reaction is
then monitored regularly by taking samples and analyzing them on SDS-PAGE or IEF gels. A fnal check is preferably done by mass spectrometry. With
some of the glycosidases like neuraminidase, one should be very careful to fully remove the enzyme (e.g. by gelfltration), since it crystallizes easily even
in minute concentrations. Another way to ensure complete removal is to use glycosidases as fusion proteins (coupled e.g. to glutathione-S-transferase; see
Grueninger-Leitch, 1996) and passing the mixture through a GST affnity column (glutathione Sepharose) after the reaction is completed.
Of course, it is always well worth a try to also crystallize the protein in its fully glycosylated form. Some proteins even crystallize better in their glycosylated
form, due to the involvement of the glycan chains in crystal contacts. Other proteins behave best when partially deglycosylated.
Glycobiology tools:
If one does get crystals from the glycosylated protein, one has to deal with handling carbohydrates, so here are a few practical tips: Modeling the ligand
into the binding site, with or without supporting electron density, of course requires a 3D structural model of the carbohydrate ligand. This can be ob-
tained either from scratch (e.g. using the modeling tool 'Sweet' from the www.glycosciences.de website, which converts carbohydrate sequences into 3D-
models) or from a structure database (via the Uppsala HIC-Up server at http://xray.bmc.uu.se/hicup/ or directly from the PDB (http://www.pdb.org/) or
other appropriate databases such as the Cambridge Structural Database (CSD; http://www.ccdc.cam.ac.uk/). The model of the carbohydrate ligand should
then be refned and checked as carefully as the protein structure itself. Currently, however, tools for analyzing carbohydrate structures are not as widely
known and applied as those for protein or DNA structures (see Kleywegt, 2003). A good tip is to check out the website at http://www.glycosciences.de. A
couple of tools, such as pdb-care (to check carbohydrate residues in pdb fles for errors) or carp (which generates Ramachandran-like plots for carbohy-
drates) or even GlyProt (to identify glycosylation sites in proteins and automatically attach them in silico) make life of structural glycobiologists signifcantly
easier. Another database, currently under development, is EuroCarbDB (http://www.eurocarbdb.org/databases). Also, questions concerning carbohydrate
nomenclature may be resolved by consulting the web site http://www.chem.qmul.ac.uk/iupac/2carb/.
by Ute Krengel, University of Oslo, September 2006.
P A G E 2 6 - 2 9
261
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P A G E 2 7 - 2 9
Sample Preparation continued...
Compound Solubility with DMSO
Compounds produced in medicinal chemistry efforts often have low aqueous solubility when compared to biological ligands. When poor compound solubil-
ity is suspected, one may try Dimethylsulfoxide (DMSO) to solubilize the compound. Dissolve the compound in 100% DMSO and THEN dilute to a lower
DMSO concentration. Dissolving a compound in dilute DMSO can be a very slow kinetic process.
Using L-Arg and L-Glu to Improve Solubility
Add 50 mM L-Arginine together with 50 mM L-Glutamic acid to the protein sample to improve protein solubility and long-term stability. Golovanov et al.
reported a 3 to 8 fold improvement in solubilization with six different test proteins.
Reference:
1. A simple method for improving protein solubility and long-term stability. Alexander P. Golovanov, Guillaume Haubergue, Stuart A. Wilson, and Lu-Yun Lian. J. Am. Chem. SOc. 2004, 126, 8933-8939.
What Sample Concentration Should I Use for My First Crystallization Screen?
For most soluble proteins, 10 to 15 mg/ml in a sample buffer that promotes the sample stability, homogeneity and monodispersity will work fne for an
initial crystallization screen.
A pre crystallization test such as the Hampton Research PCT

(HR2-140) can be used to better determine the appropriate protein concentration for crystal-
lization screening.
What to Use and What to Avoid in the Sample Buffer for Crystallization
Use the minimal number of chemicals and the minimal concentration of these chemicals necessary to promote sample stability, homogenity and
monodispersity.
Use a buffer concentration between 10 -25 mM. This will allow crystallization reagents with a buffer concentration of 50 - 100 mM to manipulate the pH of
the sample during the experiment.
Current trends seems to favor the use of 200 mM or less NaCl or KCl in the sample. Using no NaCl or KCl is just fne as well. It simply seems most overex-
pressed proteins are purifed and prepared into a fnal sample buffer contain NaCl.
If the sample is known or expected to have free sulfhydyl residues on the surface, including a reducing agent such as TCEP hydrochloride can help to
prevent oxidation of these residues. Oxidation of sulfhydryl residues can lead to sample aggregation. Avoid reducing agents if the sample has disulfde bonds
in order to prevent cleaving.
If your screens contain di or poly valent cations such as Calcium, Zinc, Iron and others, avoid Phosphate, Borate and Carbonate buffers. Such mixtures can
results in false positive salt crystals.
Phosphorylated Protein Samples
The crystallization of proteins with phosphorylation sites can often be diffcult. It is imagined that such samples are susceptible to heterogeneity due to the
presence of multiple species of protein in various states of phosphorylation. Careful sample purifcation and preparation is perhaps more of a signifcant
crystallization variable than any particular crystallization trick or specialized screen. One can use a strong anion exchange chromatography with a shallow
elution gradient to prepare a homogenous sample for crystallization. Isolation of a single phosphorylated form can produce crystals where as the presence
of multiple phosphorylated forms can lead to precipitate, microcrystals or crystals less suitable for diffraction compared to those from a homogeneous
sample. One may use mass spec to characterize the different elution samples following chromatography to assist in the selection of the best sample for
crystallization.
Batch to Batch Sample Variation
Do not mix different batches of protein. Each batch of protein should be considered and treated separately. The reason being that the expression and purif-
cation conditions as well as the chemical and physical conditions are not identical for each batch. Even subtle differences between each batch can affect the
outcome of a crystallization experiment. Each batch should be characterized and documented. Run at least an SDS-PAGE. If possible, consider also a native
PAGE, IEF-PAGE, analytical gel fltration, mass spec and dynamic light scattering to characterize each batch. Save some protein from each batch for later
experimentation, comparison, documentation, and reproducibility.
Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com 262 Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com
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Screening
Co-Crystallize
Try crystallizing your sample with a substrate, coenzyme, inhibitor, or ligand. Sometimes these agents serve to fx the macromolecule in a more compact and
stable form. This may impose a greater degree of structural homogeneity and increase the likelihood of crystallization.
Lose the Carboxy
If you cannot get diffraction quality crystals of your protein, try both Carboypeptidase-A and Carboxypeptidase-B to trim the carboxy terminus of the protein
in order to generate smaller protein fragments which might be more amicable to crystallization.
High Quality Choppers
Use sequence grade Trypsin, Chymotrypsin, SV8 Protease and subtilisin along with inhibitors to generate small, active fragments for crystal screening. (Aled
Edwards, McMaster University, CANADA & Brian McKeever, Merck, USA)
Seeing Double
Perform screens in duplicate at both 4C and room temperature. If you notice a difference in solubility between the two temperatures, be sure to include
temperature as a variable during optimization.
Screening Temperature with Limited Sample
Not enough sample to perform screens in duplicate at two different temperatures? Set up screens at room temperature. After four weeks move the plates to
4C and watch for changes in solubility. Notice a difference? Use temperature as a variable during optimization.
Screening with Volatiles
Remember when using volatile organics as the precipitating agent that no reservoir needs to be added to the drop since distillation and equilibration pro-
ceed from the reservoir to the drop.
No Crystals?
No crystals, only precipitate in initial screens? Bring precipitant/protein concentration to just below supersaturation and vary pH and temperature to ma-
nipulate sample solubility.
Low Ionic Strength
Sample soluble at low ionic strength but forms a precipitate when dialyzed against water? This "salting in" effect can be used as a method to grow crystals.
Begin with the sample in low ionic strength and dialyze the sample against a solution of lower ionic strength. Or, try changing the pH or temperature with
the sample at a constant ionic strength. Charge distribution, surface features, or conformation may change as a function of these variables which in turn may
infuence sample solubility and the ability to crystallize the sample.
Sequence Homology Search
Uncertain about the ligand requirements for your sample? If you have the amino acid sequence for your protein, consider a sequence homology search
to see if the sample is structurally related to other proteins with known ligand binding. Then try screening crystallization conditions including the ligands
identifed through the homology search.
P A G E 2 8 - 2 9
263
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P A G E 2 9 - 2 9
Screening Continued...
Going from Nano Drops to Micro Drops
If you are having trouble reproducing crystallization initially found using nano drops (100 nl plus 100 nl for example) and are having trouble growing the
crystals in the same conditions at l volumes (1 ul plus 1 ul for example), one might consider the following tips.
Heather Ringrose (Pfzer UK) has suggested that if you initially screen with 100 nl (reservoir) + 200 nl (protein) you will pick up conditions that scale up to
1 + 1 l.
Patrick Shaw-Stewart (Douglas Instruments, UK) reported that data-mining of published high-throughput results suggests that increasing the salt can help
when scaling up from nanodrops. Presumably this speeds up equilibration. Patrick suggests that if you have a hit where the main precipitant is salt, try
increasing the salt concentration in the reservoir solution by 1.0 M or more. Also, If you have a hit where the main precipitant is PEG or another organic, try
increasing reservoir salt concentration by, say, 200 mM.
Using Old Screens and Reagents
Should one use an old crystallization screen kit that is past its "Best If Used By" date, or make screen and optimization reagents with old stock solutions?
The main issue would be the ability to reproduce any crystallization results achieved with the old reagents. With time, even at 4 and -20C, the pH, ionic
strength, concentration and stability of the reagents will change. Reagents containing Polyethylene glycol undergo oxidation which produces increased lev-
els of aldehydes and peroxides which in turn lowers the pH and increases the ionic strength and produces small molecule species which can affect crystal-
lization. Evaporation from the reagents, even when frozen, increases the concentration of solutes and decreases the concentrations of solvents. The result is
that one might experience different results with an old reagent compared to a new reagent. An old reagent might not produce crystals as effectively as a new
reagent. Or, the old reagent might produce crystals not able to be grown with the new reagent. In such cases it can be diffcult or impossible to reproduce
or optimize a crystal produced with an old reagent, using freshly prepared reagents. However, in these situations, some have reported success using the old
reagent as an additive into freshly prepared reagents. For example, mix 1 part old reagent with 9 parts new reagent into the reagent well, mix and then add
this reagent to the sample drop. This procedure will incorporate some level of pH, ionic strength and small molecule change from the old reagent into the
new reagent.
The crystallization problems associated with old reagents versus new reagents seem sample-dependent. For some samples, a change in pH, ionic strength or
the presence of a small molecule contaminant produced with age may have no affect on the crystallization. For some samples, the change might be advanta-
geous and for still others, it might be other detrimental.

Biasing Crystallization Screen Reagents Towards Success
The following is a crystallization screening and optimization technique, which can be used in an attempt to improve the quality and size of a crystal or pro-
duce a different crystal form. First, one must begin with an initial crystallization reagent (ICR) which has produced some form of a crystal, microcrystalline
precipitate or similar promising result. The ICR will be used to bias a subsequent crystallization screening experiment by adding the ICR to the crystalliza-
tion drop, along with the sample and the screen reagent. Pipette the crystallization screen into the crystallization plate reservoir. One may choose to either
repeat the original screen or choose a different set of screen reagents. When creating the drop, mix the protein, screen reagent and ICR in a 3:2:1 ratio. 3
parts sample : 2 parts screen reagent : 1 part ICR. The inclusion of the ICR in the drop modifes the crystallization screen, biasing the screen reagents with a
promising reagent mix which may increase the likelihood for crystallization, improved crystals, or a different crystal form.
Possible modifcations to this technique include: (1) varying the ratio of sample : screen reagent : ICR; and (2) modifying the bias such that one uses the
ICR as the screen reagent and uses the original or new screen as the bias (3 parts protein : 2 parts ICR : 1 part screen reagent). This is essentially using the
crystallization screen as an additive screen.
References
The role of bias in crystallization conditions in automated microseeding. F. J. St John, B. Feng and E. Pozharski. Acta Cryst. (2008). D64, 1222-1227.
Enhancing protein crystallization through precipitant synergy. Majeed S, Ofek G, Belachew A, Huang CC, Zhou T, Kwong PD. Structure. 2003 Sep;11(9):1061-70.
1991-2009 Hampton Research Corp. All rights reserved. Printed in the United States of America.
This guide or parts thereof may not be reproduced in any form without the written permission of the publishers.
cryst alli zat i on
buffers t able
i n o r d e r o f p h r a n g e s
Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com 264 Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com
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d e s c r i p t i o n
Tabled below are buffers typically used in creating buffered crystallization reagents. Buffers should be used in the indicated ranges.
BUFFER 1 2 3 4 5 6 7 8 9 10 11 12
pH
1991-2009 Hampton Research Corp. All rights reserved.
Protocatechate 3,4-Dioxygenase:
New packing, old enzyme
(2.0 angstrom diffraction).
Vincent Purpero,
University of Minnesota,
Department of Biochemistry,
Molecular Biology and Biophysics
Minneapolis, MN, USA.
ACES pKa(25C) 6.8
N-(2-Acetamido)-2-aminoethanesulfonic acid
6.1 7.5
ADA pKa(25C) 6.6
N-(2-Acetamido)iminodiacetic acid
5.6 7.5
BES pKa(25C) 7.1
N,N-Bis(2-hydroxyethyl)-2-aminoethanesulfonic
acid
6.4 7.8
BIS-TRIS pKa(25C) 6.5
2,2-Bis(hydroxymethyl)-2,2',2"-nitrilotriethanol
5.8 7.2
BIS-TRIS propane pKa(25C) (1) 6.8, (2) 9.0
1,3-Bis[tris(Hydroxymethyl)methylamino]propane
6.3 9.5
Citric acid
pKa(25C) (1) 3.13, (2) 4.76, (3) 6.4
2.2 6.5
6.8 8.2
HEPES pKa(25C) 7.5
4-(2-hydroxyethyl)piperazine-1-
ethanesulfonic acid
13
HEPES sodium pKa(25C) 7.5
4-(2-Hydroxyethyl)piperazine-1-ethanesulfonic
acid sodium salt
6.6 8.5
Imidazole pKa(25C) 6.95
1,3-Diaza-2,4-cyclopentadiene, Glyoxaline
6.2 7.8
MES monohydrate pKa(25C) 6.15
2-(N-Morpholino)ethanesulfonic acid
5.2 7.1
MOPS pKa(25C) 7.2
3-(N-Morpholino)propanesulfonic acid
6.5 7.9
6.1 7.5
PIPES pKa(25C) 6.8
1,4-Piperazinediethanesulfonic acid
MOPSO pKa(25C) 6.9
b-Hydroxy-4-morpholinepropanesulfonic acid
6.2 7.6
Sodium acetate trihydrate
pKa(25C) 4.8
Acetic acid sodium salt
3.6 5.6
Sodium cacodylate trihydrate
pKa(25C) 6.2
Cacodylic acid sodium salt trihydrate
5.0 7.4
Sodium citrate tribasic dihydrate
pKa(25C) (1) 3.1, (2) 4.8, (3) 5.4
Citric acid trisodium salt dihydrate
3.0 6.2
Potassium chloride
1.0 2.2
265
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BUFFER 1 2 3 4 5 6 7 8 9 10 11 12
pH
TAPS pKa(25C) 8.4
N-[Tris(hydroxymethyl)methyl]-3-aminopropane-
sulfonic acid
7.7 9.1
TABS pKa(25C) 8.9
N-tris(Hydroxymethyl)methyl-4-aminobutane-
sulfonic acid
8.2 9.6
TAPSO pKa(25C) 7.6
2-Hydroxy-3-[tris(hydroxymethyl)methylamino]-
1-propanesulfonic acid
7.0 8.2
TEA pKa(25C) 7.8
Triethanolamine
7.3 8.3
TES pKa(25C) 7.5
N-[Tris(hydroxymethyl)methyl]-2-aminoethane-
sulfonic acid
6.8 8.2
Tricine pKa(25C) 8.1
N-[Tris(hydroxymethyl)methyl]glycine
7.4 8.8
Tris pKa(25C) 8.1
Tris(hydroxymethyl)aminomethane
7.0 9.0
7.0 9.0
TRIS hydrochloride pKa(25C) 8.1
Tris(hydroxymethyl)aminomethane
hydrochloride
13
HEPPSO pKa(25C) 7.5
N-(Hydroxyethyl)piperazine-N'-2-hydroxypro-
panesulfonic acid
7.1 8.5
6.9 8.3
MOBS pKa(25C) 7.6
4-(N-Morpholino)butanesulfonic acid
POPSO pKa(25C) 7.8
Piperazine-N,N'-bis(2-hydroxypropanesulfonic
acid)
7.2 8.5
DIPSO pKa(25C) 7.6
3-(N,N-Bis[2-hydroxyethyl]amino)-2-
hydroxypropanesulfonic acid
7.0 8.2
EPPS pKa(25C) 8.0
4-(2-Hydroxyethyl)-1-piperazinepropanesulfonic
acid; HEPPS
7.3 8.7
BICINE pKa(25C) 8.3
N,N-Bis(2-hydroxyethyl)glycine
7.4 9.3
7.6 9.0
HEPBS pKa(25C) 8.3
N-(2-Hydroxyethyl)piperazine-N'-
(4-butanesulfonic acid)
AMPSO sodium salt pKa(25C) 9.0
3-([1,1-Dimethyl-2-hydroxyethyl]amino)-2-
hydroxypropanesulfonic acid
8.3 9.7
CHES pKa(25C) 9.3
2-(Cyclohexylamino)ethanesulfonic acid
8.6 10.0
Glycine pKa(25C) 2.35
Aminoacetic acid, Aminoethanoic acid, Glycocoll
8.6 10.6
CABS pKa(25C) 10.7
4-(Cyclohexylamino)-1-butanesulfonic acid
10.0 11.4
CAPS pKa(25C) 10.4
3-(Cyclohexylamino)-1-propanesulfonic acid
9.7 11.1
CAPSO pKa(25C) 9.6
3-(Cyclohexylamino)-2-hydroxy-1-
propanesulfonic acid
8.9 10.3
Potassium chloride
12.0 13.0
266
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NH
4
Br Ammonium bromide 53.8 15
Temp (C) Reagent Formula MW
97.95
g/100 ml [M]
5.5
NH
4
Cl Ammonium chloride 26.7 20 53.49 5.0
(NH
4
)
2
C
6
H
6
O
7
Ammonium citrate dibasic 56.54 25 226.19 2.5
NH
4
F Ammonium fluoride 37.0 20 37.04 10.0
HCOONH
4
Ammonium formate 63.0 20 63.06 10.0
NH
4
I Ammonium iodide 94.2 25 144.94 6.5
NH
4
NO
3
Ammonium nitrate 90.2 25 80.04 11.2
(NH
4
)
2
HPO
4
Ammonium phosphate dibasic 46.2 25 132.06 3.5
NH
4
H
2
PO
4
Ammonium phosphate monobasic 28.7 25 115.03 2.5
(NH
4
)
2
SO
4
Ammonium sulfate 46.2 20 132.14 3.5
(NH
4
)
2
C
4
H
4
O
6
Ammonium tartrate dibasic 36.8 20 184.15 2.0
Ba(NO
3
)
2
Barium nitrate 10.2 25 261.34 0.3
CdBr
2
4H
2
O Cadmium bromide tetrahydrate 94.0 25 344.28 2.7
CdCl
2
2.5H
2
O Cadmium chloride hemipentahydrate 97.2 25 228.35 4.2
CdI
2
Cadmium iodide 73.0 20 366.22 1.9
3CdSO
4
8H
2
O Cadmium sulfate hydrate 70.3 25 769.52 0.9
CaCl
2
6H
2
O Calcium chloride hexahydrate 67.8 25 219.08 3.0
CaSO
4
2H
2
O Calcium sulfate dihydrate 0.208 25 172.17 0.01
CsBr Cesium bromide 89.8 22 212.81 4.2
CsCl Cesium chloride 126.3 25 168.36 7.5
CsI Cesium iodide 74.1 23 259.81 2.8
CsNO
3
Cesium nitrate 26.1 25 194.91 1.3
Cs
2
SO
4
Cesium sulfate 129.8 25 361.87 3.5
HOC(COOH)(CH
2
COOH)
2
H
2
O Citric acid monohydrate 88.6 25 210.14 4.2
CuBr
2
Copper(II) bromide 102.5 25 223.35 4.5
CuCl
2
2H
2
O Copper(II) chloride dihydrate 80.0 25 170.48 4.6
CuSO
4
5H
2
O Copper(II) sulfate pentahydrate 22.3 25 249.68 0.8
FeSO
4
7H
2
O Iron(III) sulfate heptahydrate 52.8 25 278.01 1.8
CH
3
COOLi 2H
2
O Lithium acetate dihydrate 51.0 20 102.02 5.0
1991-2009 Hampton Research Corp. All rights reserved.
solubi li t y t able
Face of a Lion protein crystal.
Chen, Chun-Yuan, Institute of Molecular Biology,
Academia Sinica, Taipei, Taiwan, Republic of China.
C R Y S T A L L I Z A T I O N R E A G E N T S O L U B I L I T Y
D E S C R I P T I O N
The following table provides data for formulating a saturated solution of the reagents listed at the temperature designated.
Face of a Lion protein crystal.
Chen, Chun-Yuan, Institute of Molecular Biology,
Academia Sinica, Taipei, Taiwan, Republic of China.
LiCl Lithium chloride 42.4 20
Temp (C) Reagent Formula MW
42.39
g/100 ml [M]
10.0
HOC(COOLi)(CH
2
COOLi)
2
4H
2
O Lithium citrate tribasic tetrahydrate 42.3 20 281.99 1.5
LiF Lithium fluoride 0.27 18 25.94 0.1
LiNO
3
Lithium nitrate 55.2 20 68.95 8.0
Li
2
SO
4
H
2
O Lithium sulfate monohydrate 25.6 20 127.96 2.0
MgBr
2
6H
2
O Magnesium bromide hexahydrate 83.1 18 292.20 2.8
MgCl
2
6H
2
O
C
2
H
2
O
4
MG 2H
2
O
Magnesium chloride hexahydrate
Magnesium formate dihydrate
40.7
150.04
20
25
203.30
150.38
5.0
1.0
Mg(NO
3
)
2
6H
2
O Magnesium nitrate hexahydrate 76.9 20 256.41 3.0
NiCl
2
6H
2
O Nickel(II) chloride hexahydrate 95.1 20 237.69 4.0
CH
3
COOK Potassium acetate 49.1 20 98.14 5.0
KBr Potassium bromide 56.0 25 119.00 4.7
KCl Potassium chloride 22.4 25 74.55 3.0
HOC(COOK)(CH
2
COOK)
2
H
2
O Potassium citrate tribasic monohydrate 81.2 20 324.42 2.5
KF Potassium fluoride 34.9 20 58.10 6.0
HCOOK Potassium formate 117.7 20 84.12 14.0
KI Potassium iodide 103.2 25 166.00 6.2
KNO
3
Potassium nitrate 30.3 25 101.10 3.0
K
2
HPO
4
Potassium phosphate dibasic 52.3 22 174.18 4.0
KH
2
PO
4
Potassium phosphate monobasic 20.4 22 136.09 1.5
KOCOCH(OH)CH(OH)COONa 4H
2
O Potassium sodium tartrate tetrahydrate 42.3 20 282.22 1.5
K
2
SO
4
Potassium sulfate 8.7 22 174.27 0.5
KSCN Potassium thiocyanate 77.7 22 97.18 8.0
CH
3
COONa 3H
2
O Sodium acetate trihydrate 40.8 22 136.08 3.0
NaCl Sodium chloride 29.2 22 58.44 5.0
HOC(COONa)(CH
2
COONa)
2
2H
2
O Sodium citrate tribasic dihydrate 47.1 22 294.10 1.6
NaF Sodium fluoride 3.4 22 41.99 0.8
HCOONa Sodium formate 47.6 22 68.01 7.0
NaI Sodium iodide 124.3 25 149.89 8.2
NaNO
3
Sodium nitrate 59.5 22 84.99 7.0
Na
2
HPO
4
2H
2
O Sodium phosphate dibasic dihydrate 8.9 22 177.99 0.5
NaH
2
PO
4
H
2
O Sodium phosphate monobasic monohydrate 69.0 22 137.99 5.0
Na
2
SO
4
10H
2
O Sodium sulfate decahydrate 32.2 22 322.20 1.0
C
4
H
4
Na
2
O
6
2H
2
O Sodium tartrate dibasic dihydrate 34.5 22 230.08 1.5
NaSCN Sodium thiocyanate 64.8 22 81.07 8.0
Zn(CH
3
COO)
2
2H
2
O Zinc acetate dihydrate 22.0 22 219.50 1.0
ZnSO
4
7H
2
O Zinc sulfate heptahydrate 57.5 22 287.56 2.0
1991-2009 Hampton Research Corp. All rights reserved. 267
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Jack straws.
Ulrike Demmer, Max-Planck-Institute for Biophysics, Molecular Membrane Biology, Frankfurt, Germany
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Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com 270
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ADA ........................................................... 50
Additive Screen ......................................... 28
Additive Screen HT ................................... 28
Adjustable Crystal Mount ...............145, 152
Adjustable Mounted CryoLoops ............. 125
Agarose .............................................57, 104
Als Oil ....................................................... 56
ALS Style CrystalCap ............................. 121
AlumaSeal II Sealing Film ...................... 101
Ammonium acetate ................................... 43
Ammonium chloride .................................. 43
Ammonium citrate dibasic ........................ 43
Ammonium fluoride ................................... 43
Ammonium formate .................................. 43
Ammonium nitrate ..................................... 43
Ammonium phosphate dibasic ................. 43
Ammonium phosphate monobasic ........... 43
Ammonium sulfate .................................... 44
Ammonium tartrate dibasic ....................... 44
Applicators .............................................. 103
Batch Crystallization ......................79, 87-89
Beeswax .................................................. 143
BICINE ...................................................... 50
BIS-TRIS ................................................... 50
BIS-TRIS propane ................................... 50
Books ...............................................166-173
Brass Pin .........................................145, 152
Brass Pin with Platform ..................145, 152
Buffer Kits. ............................................63-67
Buffer Reagents ...................................50-52
Cadmium chloride hydrate........................ 44
Cadmium sulfate hydrate .......................... 44
Calcium acetate hydrate ........................... 44
Calcium chloride dihydrate ....................... 44
Canned Air - Duster .................................. 95
Cap Mat for MASTERBLOCK Plate ....... 101
Capillaries ........................................138-140
Capillary Cutting Stone .......................... 140
Capillary Supplies ...........................138-145
Capillary Support .............................138-145
Capillary Wax & Clay ......................143-144
Cesium chloride ........................................ 44
Chelating Resin ....................................... 115
Circle Cover Slides ..............................91-93
Citric acid .................................................. 50
Citric acid BIS-TRIS propane ................... 50
Clamp ...................................................... 131
Clay ......................................................... 144
ClearSeal Film .......................................... 99
ClearSeal Film Applicator ......................... 99
Cobalt(II) chloride hexahydrate ................ 44
Containerless Crystallization .................... 56
Corning CrystalEX 96 Well Plates .......81-82
Corning CrystalEX 384 Well Plates .......... 90
Cover Slides ........................................91-95
Cover Slide Dispensers ............................ 92
Cover Slide Holder .............................92, 94
Cover Slide Vacuum Gadget .................... 94
Cryo Labels ......................................161,163
CryoCane Color Coders ......................... 133
CryoCanes .............................................. 133
Cryocrystallography ......................... 118-135
CryoLoops........................................124-127
CryoPro Kit ................................................ 68
Cryoprotectants ...................................54, 68
CryoSleeves ............................................ 133
CryoTongs ........................................128-129
Cryschem 24-1 SBS Plate ........................ 72
Cryschem Plate......................................... 72
Crystal Clear Sealing Film ........................ 99
Crystal Clear Sealing Tape ..................... 100
Crystal Dye - Izit ....................................... 58
Crystal Pencil .......................................... 113
Crystal Probe .......................................... 113
Crystal Screen 2 Kit .................................8-9
Crystal Screen Cryo Kit .......................22-23
Crystal Screen HT ...................................8-9
Crystal Screen Kit ....................................8-9
Crystal Screen Lite Kit .........................18-19
CrystalBridge ............................................. 76
CrystalCap Colored ................................ 123
CrystalCap Copper ................................. 123
CrystalCap Copper Magnetic ALS HT ... 119
CrystalCap Copper Magnetic HT ........... 118
CrystalCap Ex ......................................... 122
CrystalCap Holder................................... 132
CrystalCap HT Systems .................. 118-120
CrystalCap Magnetic .............................. 121
CrystalCap Magnetic Systems ........121-122
CrystalCap Systems ........................122-123
CrystalClear Strips .................................... 80
CrystalDrop Lids ....................................... 83
CrystalEX 96 Well Plates .....................81-82
CrystalEX 384 Well Plates........................ 90
Crystallization Plates & Accessories .72-104
CrystalQuick Plates .................................. 83
CrystalWands .......................................... 130
CrystalWand Magnetic ............................ 130
CTAB ......................................................... 44
Custom Shop .......................................36-37
Cutting Stone .......................................... 140
Deep Well MASTERBLOCK..................... 86
Detergent Screen ...................................... 29
Detergent Screen HT ................................ 29
Dewar Flasks ...................................134-135
Dialysis Button Applicators ..................... 103
Dialysis Buttons ...................................... 102
Dialysis Membrane Discs ....................... 103
1,4-Dioxane ............................................... 42
DL-Malic acid ............................................ 46
DMS Oil ..................................................... 56
Douglas Instruments Plates................80, 89
Dow Corning 7 Release Compound ........ 98
Dow Corning Vacuum Grease .................. 98
Duco Cement .......................................... 142
Dye ............................................................ 58
Epoxy ...................................... 126-127, 142
Ethylene glycol .......................................... 42
Ethylene imine polymer ............................ 40
Fluorinert ................................................... 55
FMS Oil ..................................................... 56
Foam Dewars.......................................... 135
Forceps ............................................ 110-113
Gels ........................................................... 57
Gizmo - Cover Slide Dispenser ................ 92
Glass Capillaries ..............................138-139
Glass Cover Slides ..............................91-93
Glass Fibers ............................................ 140
Glass Number 50 Capillaries .......... 138-139
Glass Sitting Drop Rods ........................... 95
Glucose Isomerase ................................. 176
Glue ......................................... 126-127, 142
Glycerol ..................................................... 42
Goniometer Heads & Supplies ........148-153
Granada Crystallization Box ................... 104
Grease ...................................................... 98
Grease Cartridges .................................... 98
Greiner 24 Well ComboPlate .................... 76
Greiner 96 Well Imp@ct Plates ................ 88
Greiner CrystalBridge ............................... 76
Greiner CrystalDrop Lid ............................ 83
Greiner CrystalQuick Plates ..................... 83
Greiner Microbatch ................................... 79
Greiner Plate Lids ..................................... 88
Grid Screens ........................................14-15
Grid Screen Ammonium Sulfate
Grid Screen MPD
Grid Screen PEG 6000
Grid Screen PEG/LiCl
Grid Screen Salt HT
Grid Screen Sodium Chloride
Grid Screen Sodium Malonate
Heavy Atom Screens ...........................30-32
HEPES ...................................................... 50
HEPES sodium ......................................... 50
1,6-Hexanediol .......................................... 42
Hydrochloric acid ...................................... 52
Hydrogel .................................................... 57
I3C Phasing Kit ......................................... 32
Imidazole ................................................... 51
Immersion Oil ............................................ 97
Imp@ct Plates........................................... 88
Index HT ...................................................6-7
Index Kit ...................................................6-7
Individual Screen Reagents.................36-37
Intelli-Plates ...................................73, 78, 84
Intelli-Plate 24-4 well ................. 73
Intelli-Plate 48-2 well ................. 78
Intelli-Plate 48-3 well ................. 78
Intelli-Plate 96 Flat Shelf ........... 84
Intelli-Plate 96-2 LVR ................ 84
Intelli-Plate 96-2 Original ........... 84
Intelli-Plate 96-3 well ................. 84
Iron(III) chloride hexahydrate .................... 44
Izit - Crystal Dye ....................................... 58
Jeffamine ................................................... 40
L-Proline .................................................... 47
Labels & Pens ..................................160-163
Linbro Plate ............................................... 76
Lipase B .................................................. 179
Lithium acetate dihydrate ......................... 45
Lithium chloride ......................................... 45
Lithium citrate tribasic tetrahydrate .......... 45
Lithium nitrate............................................ 45
Lithium sulfate monohydrate .................... 45
LM Agarose .......................................57, 104
Long CryoTongs ..................................... 129
Long CrystalWands ................................ 130
Low Form Dewar .................................... 134
Low Ionic Strength Screen Kit .................. 25
Lysozyme ................................................ 177
Lysozyme Crystallization Reagent ......... 177
Magnesium acetate tetrahydrate .............. 45
Magnesium chloride hexahydrate ............ 45
Magnesium formate dihydrate .................. 45
Magnesium nitrate hexahydrate ............... 45
Magnesium sulfate heptahydrate ............. 45
Magnesium sulfate hydrate ...................... 46
Magnetic Bases ...................................... 153
Malic acid .................................................. 46
MASTERBLOCK Deep Well Plate ........... 86
MembFac HT .......................................18-19
MembFac Kit ........................................18-19
MES monohydrate .................................... 51
Micro-Bridges ............................................ 96
Micro-Bridges Polypropylene .................... 96
Micro-Tools .......................................108-109
Microbatch Plates ..........................79, 87-89
Microdialysis. ....................................102-103
Microdialysis Buttons .............................. 102
Microdialysis Membrane Discs ............... 103
MicroTubes.............................................. 126
MicroWick ................................................ 141
Modular VDX Plates ................................. 77
Modular VDX Plates with sealant ............. 77
Mounted CryoLoops ........................124-125
Mounting Clay - Four Color .................... 144
MPD .......................................................... 42
MRC Plates ..........................................85-87
A
B
C
D
E
F
G
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MRC Plate Seal ...................................... 100
Multichannel Pipetter Basin ...................... 90
Natrix Kit ...............................................20-21
NDSB ........................................................ 52
Nickel(II) chloride hexahydrate ................. 46
Nucleic Acid Mini Screen Kit ..................... 24
Oils .......................................................55-56
OptiClear Cover Slides ............................. 93
Optimize Reagents ..............................40-58
Organic (Non-Volatile) Reagents .............. 42
Organic (Volatile) Reagents ...................... 42
Paper Wicks ............................................ 141
Paraffin Oil. ............................................... 56
Paratone-N. ............................................... 54
PCT Kit ........................................................ 5
PEG ......................................................40-42
PEG/Ion 2 Screen Kit ..........................12-13
PEG/Ion Screen Kit..............................12-13
PEG/Ion HT ..........................................12-13
PEGRx 1 Kit ......................................... 10-11
PEGRx 2 Kit ......................................... 10-11
PEGRx HT ........................................... 10-11
PEN-VAC Kit ............................................. 94
Pencil ....................................................... 113
Pens ........................................................ 160
Pentaerythritol ........................................... 40
Perfluoropolyether ..................................... 54
pH Kits ..................................................63-67
Plain Cover Slides .................................... 93
Plastic Cover Slides .................................. 93
Plate Lids ............................................83, 88
Plate Stand................................................ 94
Plates & Accessories .........................72-104
9 Well Plate .......................................... 97
24 Well Plates .................................72-77
48 Well Plates .................................78-79
72 Well Plates ...................................... 79
96 Well Plates .................................80-90
384 Well Plate ...................................... 90
Poly(acrylic acid sodium salt) 5,100 ......... 40
Polyethylene glycol ..............................40-42
Polymer Reagents. ..............................40-42
Polypropylene glycol P 400 ...................... 42
Polyvinylpyrrolidone K 15 ......................... 42
Potassium acetate .................................... 46
Potassium bromide ................................... 46
Potassium chloride ................................... 46
Potassium citrate tribasic monohydrate ... 46
Potassium fluoride .................................... 46
Potassium formate .................................... 46
Potassium phosphate dibasic .............46, 48
Potassium phosphate monobasic ............ 47
Potassium nitrate ...................................... 47
Potassium sodium tartrate tetrahydrate ... 47
Potassium sulfate...................................... 47
Potassium thiocyanate .............................. 47
Pre-Crystallization Test ............................... 5
Proline ....................................................... 47
2-Propanol ................................................. 42
Protein Standards ............................176-179
Quartz Capillaries ............................138-139
Quik Optimize Kit ...................................... 48
Quik Screen Kit ....................................14-15
Reagents ..............................................40-58
Red Sticky Wax ....................................... 144
Reducing Agent......................................... 53
Robolid .................................................... 101
Round Capillaries.................................... 104
Salt Kit ....................................................... 62
Salt Reagents ......................................43-49
SaltRx 1 Kit ..........................................16-17
SaltRx 2 Kit ..........................................16-17
SaltRx HT .............................................16-17
Sandwich Box Setup ................................ 97
Screens ..................................................5-32
Sealants ...............................................97-98
Sealing Film Applicator ............................. 99
Sealing Films .....................................99-101
Sealing Tapes ......................................... 100
Seed Bead Kit ......................................... 114
Seeding ............................................ 114-115
Seeding Tool ........................................... 115
Set Screws .............................................. 151
Silica Hydrogel .......................................... 57
Silicon Oil .................................................. 56
Siliconized Cover Slides ........................... 91
Silver Bullets Bio HT ............................26-27
Silver Bullets Bio Kit .............................26-27
Silver Bullets HT ..................................26-27
Silver Bullets Kit ...................................26-27
Sitting Drop Rods ...................................... 95
Slide Dispensers ....................................... 92
Slides ....................................................91-95
Sodium acetate trihydrate ...................47, 51
Sodium bromide ........................................ 47
Sodium cacodylate trihydrate ................... 51
Sodium chloride ........................................ 47
Sodium citrate tribasic dihydrate ........48, 51
Sodium fluoride ......................................... 48
Sodium formate......................................... 48
Sodium hydroxide ..................................... 52
Sodium malonate ...................................... 48
Sodium nitrate ........................................... 48
Sodium phosphate dibasic dihydrate ....... 48
Sodium phosphate monobasic monohyd . 48
Sodium potassium phosphate .................. 48
Sodium sulfate decahydrate ..................... 48
Sodium tartrate dibasic dihydrate ............. 49
Sodium thiocyanate .................................. 49
Solubilizing Agents .................................... 52
Solvent Resistant Pens .......................... 160
Special Glass 10 Capillaries ........... 138-139
Square Cover Slides ............................91-93
Stereo Viewer ......................................... 173
Stereopticon ............................................ 173
StockOptions Buffer Kits ......................64-67
StockOptions Bis-Tris Kit ..................... 66
StockOptions Citric Acid ...................... 64
StockOptions HEPES Kit ..................... 66
StockOptions MES Kit ......................... 65
StockOptions Sodium Acetate Kit ....... 64
StockOptions Sodium Cacodylate Kit . 65
StockOptions Sodium Citrate Kit ........ 65
StockOptions Sodium HEPES Kit ....... 66
StockOptions Tris Kit ........................... 67
StockOptions Tris Hydrochloride Kit ... 67
StockOptions CryoPro Kit ......................... 68
StockOptions Kits.................................62-68
StockOptions pH Kit.................................. 63
StockOptions Salt Kit ................................ 62
Succinic acid ............................................. 49
Super Glue .............................. 126-127, 142
Tacsimate .................................................. 49
Tapes ....................................................... 100
TCEP hydrochloride .................................. 53
Trimethylamine N-oxide dihydrate ............ 49
Tris ............................................................. 51
TRIS hydrochloride ................................... 52
Tryptone .................................................... 49
Tube Clamp ............................................. 131
UVP Hanging Drop MRC Plate Seal ...... 100
Vacuum Gadget ........................................ 94
Vacuum Grease ........................................ 98
Vapor Batch Plates ................................... 89
VDX48 Well Plate with sealant ................. 79
VDX Plate.................................................. 74
VDX Plate with sealant ............................. 74
VDX Plate - Modular ................................. 77
VDXm Plate .............................................. 75
VDXm Plate with sealant .......................... 75
Vial Clamps ............................................. 131
Viewers .................................................... 173
Wax Pen .................................................. 143
Waxes ..............................................143-144
Wicks ....................................................... 141
Xenon Chamber & Accessories ............. 156
Xenon Derivatization ........................156-157
Xenon Recovery System ........................ 157
Xylanase ................................................. 178
XYZ Heated Goniometer Heads .....148-149
XYZ Standard Goniometer Heads ..150-152
Z Capillaries ............................................ 152
Z Platforms .............................................. 153
Zinc acetate dihydrate .............................. 49
Zinc chloride .............................................. 49
Zinc sulfate heptahydrate ......................... 49
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HR1-002 ................................................. 72
HR2-078 ............................................ 26-27
HR2-080 ............................................ 26-27
HR2-082 ............................................ 10-11
HR2-084 ............................................ 10-11
HR2-086 ............................................ 10-11
HR2-088 ............................................ 26-27
HR2-090 ............................................ 26-27
HR2-092 ............................................ 26-27
HR2-094 ............................................ 26-27
HR2-096 ............................................ 26-27
HR2-098 ............................................ 12-13
HR2-100 ................................................. 67
HR2-102 ................................................. 66
HR2-104 ................................................. 64
HR2-106 ................................................. 66
HR2-107 ............................................ 16-17
HR2-109 ............................................ 16-17
HR2-110 ................................................. 8-9
HR2-112 ................................................. 8-9
HR2-114 ............................................. 18-19
HR2-116 ............................................. 20-21
HR2-118 ...................................................24
HR2-120 ................................................. 25
HR2-122 ............................................ 22-23
HR2-126 ............................................ 12-13
HR2-128 ............................................ 18-19
HR2-130 ................................................ 8-9
HR2-132 ................................................. 68
HR2-134 ................................................ 6-7
HR2-136 ............................................ 16-17
HR2-137 ............................................ 18-19
HR2-138 ................................................. 28
HR2-139 ............................................ 12-13
HR2-140 ................................................... 5
HR2-142 ................................................... 5
HR2-144 ................................................ 6-7
HR2-211 ............................................. 14-15
HR2-213 ............................................ 14-15
HR2-215 ............................................ 14-15
HR2-217 ............................................ 14-15
HR2-219 .................................... 14-15, 177
HR2-221 ............................................ 14-15
HR2-223 ................................................. 48
HR2-231 ................................................. 66
HR2-233 ......................................... 64, 177
HR2-235 ................................................. 65
HR2-237 ................................................. 67
HR2-239 ................................................. 65
HR2-241 ................................................. 63
HR2-243 ................................................. 65
HR2-245 ................................................. 62
HR2-247 ............................................ 14-15
HR2-248 ............................................ 14-15
HR2-310 ................................................. 57
HR2-312 ............................................... 115
HR2-320 ............................................... 114
HR2-406 ........................................... 29, 36
HR2-408 ................................................. 29
HR2-409 ................................................. 39
HR2-428 ........................................... 28, 36
HR2-442 ...................................... 30-31, 36
HR2-444 ...................................... 30-31, 36
HR2-446 ...................................... 30-31, 36
HR2-448 ...................................... 30-31, 36
HR2-450 ...................................... 30-31, 36
HR2-501 ................................................. 40
HR2-503 ................................................. 40
HR2-507 ................................................. 50
HR2-509 ................................................. 50
HR2-513 ................................................. 41
HR2-515 ................................................. 41
HR2-517 ................................................. 41
HR2-523 ................................................. 41
HR2-525 ................................................. 41
HR2-527 ................................................. 41
HR2-529 ................................................. 41
HR2-533 ................................................. 41
HR2-535 ................................................. 41
HR2-537 ................................................. 45
HR2-539 ................................................. 47
HR2-541 ................................................. 44
HR2-543 ................................................. 47
HR2-545 ................................................. 45
HR2-547 ................................................. 48
HR2-549 ................................................. 48
HR2-551 ................................................. 48
HR2-553 ................................................. 47
HR2-555 ................................................. 43
HR2-557 ................................................. 44
HR2-559 ................................................. 45
HR2-561 ................................................. 45
HR2-563 ................................................. 49
HR2-565 ................................................. 43
HR2-567 ................................................. 44
HR2-569 ................................................. 51
HR2-571 ................................................. 51
HR2-573 ................................................. 51
HR2-575 ................................................. 51
HR2-577 ................................................. 50
HR2-579 ................................................. 52
HR2-581 ................................................. 52
HR2-583 ................................................. 52
HR2-585 ................................................. 50
HR2-587 ................................................. 51
HR2-589 ................................................. 51
HR2-591 ................................................. 41
HR2-593 ................................................. 56
HR2-595 ................................................. 56
HR2-597 ................................................. 40
HR2-599 ................................................. 40
HR2-601 ................................................. 40
HR2-603 ................................................. 41
HR2-605 ................................................. 41
HR2-607 ................................................. 41
HR2-609 ................................................. 41
HR2-611 ...................................................41
HR2-613 ................................................. 41
HR2-615 ................................................. 42
HR2-617 ................................................. 42
HR2-619 ................................................. 42
HR2-621 ................................................. 42
HR2-623 ................................................. 42
HR2-625 ................................................. 42
HR2-627 ................................................. 42
HR2-629 ................................................. 43
HR2-631 ................................................. 45
HR2-633 ................................................. 46
HR2-635 ........................................... 46, 48
HR2-637 ......................................... 47, 177
HR2-639 ................................................. 48
HR2-641 ................................................. 49
HR2-643 ................................................. 54
HR2-645 ................................................. 48
HR2-647 ................................................. 46
HR2-649 ................................................. 46
HR2-651 ................................................. 53
HR2-657 ................................................. 45
HR2-659 ................................................. 43
HR2-661 ................................................. 48
HR2-663 ................................................. 47
HR2-665 ................................................. 43
HR2-667 ................................................. 46
HR2-669 ................................................. 45
HR2-671 ................................................. 46
HR2-673 ................................................. 48
HR2-675 ................................................. 47
HR2-677 ................................................. 49
HR2-679 ................................................. 44
HR2-681 ................................................. 45
HR2-683 ................................................. 46
HR2-685 ................................................. 43
HR2-687 ................................................. 46
HR2-689 ................................................. 43
HR2-691 ................................................. 43
HR2-693 ................................................. 49
HR2-695 ................................................. 47
HR2-697 ................................................. 45
HR2-699 ................................................. 47
HR2-701 ................................................. 52
HR2-703 ................................................. 52
HR2-705 ................................................. 52
HR2-707 ................................................. 48
HR2-709 ................................................. 49
HR2-711 ...................................................44
HR2-713 ................................................. 44
HR2-715 ................................................. 44
HR2-717 ................................................. 44
HR2-719 ................................................. 44
HR2-721 ................................................. 44
HR2-723 ................................................. 50
HR2-725 ................................................. 51
HR2-727 ................................................. 52
HR2-729 ................................................. 50
HR2-731 ......................................... 51, 177
HR2-733 ................................................. 50
HR2-735 ................................................. 51
HR2-737 ................................................. 51
HR2-739 ................................................. 40
HR2-741 ................................................. 40
HR2-743 ................................................. 40
HR2-745 ................................................. 40
HR2-747 ................................................. 48
HR2-749 ................................................. 48
HR2-751 ................................................. 48
HR2-755. ................................................ 49
HR2-757 ................................................. 50
HR2-759 ................................................. 43
HR2-761 ................................................. 46
HR2-763 ................................................. 47
HR2-765 ................................................. 48
HR2-767 ................................................. 44
HR2-769 ................................................. 42
HR2-771 ................................................. 42
HR2-773 ................................................. 40
HR2-775 ................................................. 47
HR2-777 ................................................. 49
HR2-779 ................................................. 46
HR2-781 ................................................. 50
HR2-783 ................................................. 50
HR2-785 ................................................. 50
HR2-787 ................................................. 51
HR2-789 ................................................. 51
HR2-791 ................................................. 52
HR2-793 ................................................. 52
HR2-795 ................................................. 50
HR2-797 ................................................. 55
HR2-799 ................................................. 43
HR2-801 ................................................. 53
HR2-803 ................................................. 45
HR2-805 ............................................... 177
H R 1 -
H R 2 -
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HR2-807 ................................................. 48
HR2-809 ................................................. 48
HR2-811 ................................................. 49
HR2-813 ................................................. 49
HR2-814 ................................................. 54
HR2-817 ................................................. 50
HR2-819 ................................................. 51
HR2-821 ................................................. 45
HR2-823 ................................................. 49
HR2-825 ................................................. 49
HR2-827 ................................................. 49
HR2-829 ................................................. 49
HR2-831 ................................................. 50
HR2-833 ................................................. 50
HR2-835 ................................................. 49
HR2-837 ................................................. 41
HR2-839 ................................................. 49
HR2-841 ................................................. 41
HR2-843 ................................................. 49
HR2-845 ................................................. 41
HR2-847 ................................................. 49
HR2-849 ................................................. 41
HR2-851 ................................................. 49
HR2-900-** ....................................... 37, 67
HR2-902-** ....................................... 37, 66
HR2-904-** ....................................... 37, 64
HR2-906-** ....................................... 37, 66
HR2-906-24 ............................................ 50
HR2-907-** .................................. 16-17, 36
HR2-909-** .................................. 16-17, 36
HR2-910-** ...................................... 8-9, 36
HR2-912-** ...................................... 8-9, 36
HR2-914-** .................................. 22-23, 36
HR2-916-** .................................. 18-19, 36
HR2-918-** .................................. 20-21, 36
HR2-920-** .................................. 18-19, 36
HR2-921-** .................................. 14-15, 36
HR2-922-** .................................. 12-13, 36
HR2-924-** .................................. 14-15, 36
HR2-926-** .................................. 14-15, 36
HR2-928-** .................................. 14-15, 36
HR2-930-** .................................. 14-15, 36
HR2-931-** ....................................... 37, 66
HR2-931-01 ............................................ 50
HR2-932-** .................................. 14-15, 36
HR2-933-** ....................................... 37, 64
HR2-935-** ....................................... 37, 65
HR2-937-** ....................................... 37, 67
HR2-939-** ....................................... 37, 65
HR2-940-** ......................................... 5, 36
HR2-941-** ....................................... 37, 63
HR2-943-** ....................................... 37, 65
HR2-943-07 ............................................ 51
HR2-944-** ...................................... 6-7, 36
HR2-947-** .................................. 14-15, 36
HR2-982-** .................................. 10-11, 36
HR2-984-** .................................. 10-11, 36
HR2-988-** .................................. 26-27, 36
HR2-993-** ............................................. 37
HR2-995-** ............................................. 37
HR2-996-** .................................. 26-27, 36
HR2-997-** ............................................. 37
HR2-998-** .................................. 12-13, 36
HR2-999-** ............................................. 37
HR3-082 ................................................. 85
HR3-083 ................................................. 85
HR3-084 ................................................. 88
HR3-085 ................................................. 88
HR3-086 ................................................. 79
HR3-087 ................................................. 79
HR3-088 ................................................. 83
HR3-089 ................................................. 83
HR3-090 ................................................. 83
HR3-091 ................................................. 83
HR3-092G .............................................. 83
HR3-093G .............................................. 83
HR3-094G .............................................. 83
HR3-095G .............................................. 83
HR3-096 ................................................. 83
HR3-097 ................................................. 83
HR3-098 ................................................. 88
HR3-099 ................................................. 88
HR3-100 ................................................. 88
HR3-101 ................................................. 88
HR3-102 ................................................. 87
HR3-103 ................................................. 87
HR3-104 ............................................... 104
HR3-105 ................................................. 86
HR3-108 ................................................. 75
HR3-110 ................................................. 76
HR3-111..................................................101
HR3-112 ................................................. 76
HR3-113 ................................................. 81
HR3-114 ................................................. 73
HR3-115 ................................................. 81
HR3-120 ................................................. 79
HR3-122 ................................................. 79
HR3-123 ................................................. 86
HR3-125 ................................................. 86
HR3-128 ................................................. 80
HR3-132 ................................................. 97
HR3-133.................................................... 32
HR3-134 ................................................. 97
HR3-136 ................................................. 97
HR3-137 ................................................. 83
HR3-138 ................................................. 83
HR3-140 ................................................. 74
HR3-142 ................................................. 74
HR3-143 ................................................. 84
HR3-145 ................................................. 84
HR3-146 ................................................. 95
HR3-148 ................................................. 95
HR3-150 ................................................. 76
HR3-154 ................................................. 76
HR3-158 ................................................. 72
HR3-160 ................................................. 72
HR3-162 ................................................. 80
HR3-170 ................................................. 74
HR3-172 ................................................. 74
HR3-183 ................................................. 84
HR3-185 ................................................. 84
HR3-190 ................................................. 83
HR3-192 ................................................. 83
HR3-194 ............................................... 104
HR3-196 ................................................. 77
HR3-198 ................................................. 77
HR3-200 ................................................. 76
HR3-202 ................................................. 98
HR3-204 ................................................. 77
HR3-207T ............................................... 93
HR3-209T ............................................... 93
HR3-211 ...................................................93
HR3-213 ................................................. 93
HR3-215 ................................................. 91
HR3-217 ................................................. 91
HR3-219 ................................................. 93
HR3-221 ................................................. 93
HR3-223 ................................................. 91
HR3-225 ................................................. 91
HR3-227 ................................................. 93
HR3-229 ................................................. 93
HR3-231 ................................................. 91
HR3-233 ................................................. 91
HR3-235 ................................................. 93
HR3-237 ................................................. 93
HR3-239 ................................................. 91
HR3-241 ................................................. 91
HR3-243 ................................................. 93
HR3-245 ................................................. 93
HR3-247 ................................................. 91
HR3-249 ................................................. 91
HR3-251 ................................................. 94
HR3-265 ................................................. 89
HR3-267 ................................................. 89
HR3-269 ................................................. 90
HR3-271 ................................................. 81
HR3-273 ................................................. 81
HR3-275 ................................................. 79
HR3-277 ................................................. 91
HR3-278T ............................................... 91
HR3-279 ................................................. 91
HR3-280T ............................................... 91
HR3-281 ................................................. 83
HR3-283 ................................................. 83
HR3-284 ................................................. 83
HR3-285 ................................................. 83
HR3-293 ................................................. 88
HR3-295 ................................................. 88
HR3-297 ................................................. 84
HR3-299 ................................................. 84
HR3-302 ................................................. 83
HR3-304 ................................................. 83
HR3-306 ................................................. 75
HR3-310 ................................................. 96
HR3-312 ................................................. 96
HR3-314 ............................................... 102
HR3-316 ............................................... 102
HR3-318. .............................................. 102
HR3-320 ............................................... 102
HR3-322 ............................................... 102
HR3-326 ............................................... 102
HR3-328 ............................................... 102
HR3-330 ............................................... 102
HR3-332 ............................................... 102
HR3-336 ............................................... 102
HR3-338 ............................................... 103
HR3-340 ................................................. 96
HR3-342 ................................................. 96
HR3-344 ............................................... 103
HR3-346 ............................................... 103
HR3-411 ...................................................56
HR3-413 ................................................. 56
HR3-415 ................................................. 56
HR3-417 ................................................. 56
HR3-421 ................................................. 56
HR3-423 ................................................. 56
HR3-508 ................................................. 98
HR3-510 ................................................. 98
HR3-511 ............................................... 100
HR3-515 ................................................. 91
HR3-517 ................................................. 91
HR3-607 ............................................... 100
HR3-609 ................................................. 99
HR3-611 ................................................. 97
HR3-613 ................................................. 97
HR3-615 ................................................. 97
HR3-617 ................................................. 97
HR4-110 .................................................141
HR4-112 .................................................141
HR4-114 .................................................141
H R 4 -
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Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com 274
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HR4-116 .................................................141
HR4-122 ............................................... 141
HR4-211 .................................................141
HR4-213 ............................................... 141
HR4-217 ............................................... 113
HR4-310 ............................................... 144
HR4-312 ............................................... 143
HR4-313 ............................................... 124
HR4-315 ............................................... 130
HR4-316 ................................ 126-127, 142
HR4-318 ................................ 126-127, 142
HR4-320 ............................................... 142
HR4-326 ............................................... 144
HR4-328 ............................................... 143
HR4-330 ............................................... 141
HR4-334 ............................................... 140
HR4-336 ............................................... 125
HR4-338 ............................................... 125
HR4-342 ............................................... 143
HR4-344 ............................................... 143
HR4-346 ................................ 126-127, 142
HR4-348.................................................. 103
HR4-350.................................................. 103
HR4-400 ............................................... 163
HR4-402 ............................................... 163
HR4-404 ............................................... 161
HR4-406 ............................................... 161
HR4-411 ...................................................95
HR4-436 ............................................... 162
HR4-438 ............................................... 162
HR4-440 ............................................... 162
HR4-442 ............................................... 162
HR4-444 ............................................... 162
HR4-446 ............................................... 162
HR4-456 ............................................... 163
HR4-458 ............................................... 160
HR4-460 ............................................... 161
HR4-462 ............................................... 160
HR4-464 ............................................... 160
HR4-466 ............................................... 160
HR4-468 ................................................. 94
HR4-506 ............................................... 100
HR4-511 ............................................... 100
HR4-513 ............................................... 173
HR4-515 ............................................... 173
HR4-517 ............................................... 173
HR4-521 ................................................. 99
HR4-523 ................................................. 99
HR4-525 ......................................... 99, 101
HR4-600 ............................................... 130
HR4-602 ............................................... 130
HR4-615 ............................................... 124
HR4-617 ............................................... 124
HR4-619 ............................................... 130
HR4-623 ............................................... 126
HR4-625 ............................................... 124
HR4-627 ............................................... 153
HR4-629 ............................................... 153
HR4-631 ............................................... 128
HR4-633 ............................................... 128
HR4-635 ............................................... 128
HR4-637 ....................................... 120, 128
HR4-639 ............................................... 128
HR4-641 ............................................... 128
HR4-643 ............................................... 150
HR4-647 ............................................... 150
HR4-651 ............................................... 153
HR4-653 ............................................... 153
HR4-657 ............................................... 149
HR4-659 ............................................... 151
HR4-661 ....................................... 145, 152
HR4-663 ....................................... 145, 152
HR4-665 ............................................... 123
HR4-667 ............................................... 128
HR4-669 ............................................... 149
HR4-670 ............................................... 131
HR4-671 ............................................... 131
HR4-672 ............................................... 131
HR4-673 ............................................... 135
HR4-675 ............................................... 135
HR4-677 ............................................... 104
HR4-678 ............................................... 104
HR4-679 ............................................... 104
HR4-680 ............................................... 104
HR4-681 ............................................... 104
HR4-682 ............................................... 104
HR4-683 ............................................... 104
HR4-684 ............................................... 104
HR4-685 ............................................... 104
HR4-686 ............................................... 104
HR4-687 ............................................... 104
HR4-688 ............................................... 104
HR4-695 ............................................... 134
HR4-697 ............................................... 134
HR4-699 ............................................... 134
HR4-701 ............................................... 134
HR4-705 ............................................... 132
HR4-706 ............................................... 132
HR4-707 ............................................... 132
HR4-708 ............................................... 133
HR4-709 ............................................... 133
HR4-710 ................................................. 58
HR4-711 ............................................... 133
HR4-713 ............................................... 133
HR4-715 ............................................... 133
HR4-717 ............................................... 133
HR4-719 ............................................... 133
HR4-721 ............................................... 133
HR4-722 ............................................... 133
HR4-727 ............................................... 131
HR4-729 ............................................... 130
HR4-731 ............................................... 121
HR4-733 ............................................... 121
HR4-737 ............................................... 122
HR4-739 ............................................... 122
HR4-741 ............................................... 122
HR4-743 ............................................... 122
HR4-745 ............................................... 122
HR4-747 ............................................... 122
HR4-749 ............................................... 122
HR4-753 ............................................... 151
HR4-755 ............................................... 149
HR4-761 ............................................... 153
HR4-763 ............................................... 153
HR4-765 ............................................... 148
HR4-767 ............................................... 148
HR4-769 ............................................... 152
HR4-771 ............................................... 152
HR4-775 ............................................... 149
HR4-777 ............................................... 149
HR4-779 ............................................... 121
HR4-781 ............................................... 156
HR4-791 ............................................... 156
HR4-793 ............................................... 156
HR4-795 ............................................... 156
HR4-797 ............................................... 157
HR4-799 ............................................... 156
HR4-811 .................................................108
HR4-813 ............................................... 108
HR4-815 ............................................... 108
HR4-817 ............................................... 108
HR4-819 ............................................... 108
HR4-821 ............................................... 108
HR4-823 ............................................... 108
HR4-825 ............................................... 108
HR4-827 ............................................... 108
HR4-829 ....................................... 108, 109
HR4-831 ....................................... 108, 109
HR4-835 ............................................... 113
HR4-837 ............................................... 109
HR4-839 ............................................... 109
HR4-841 ............................................... 109
HR4-843 ............................................... 109
HR4-845 ............................................... 109
HR4-847 ............................................... 109
HR4-849 ............................................... 109
HR4-851 ............................................... 109
HR4-853 ............................................... 109
HR4-855 ............................................... 110
HR4-857 ............................................... 110
HR4-859 ............................................... 110
HR4-861 ............................................... 110
HR4-863 ................................................111
HR4-865 ................................................111
HR4-869 ................................................111
HR4-871 ............................................... 112
HR4-875 ............................................... 112
HR4-879 ............................................... 112
HR4-881 ............................................... 112
HR4-883 ............................................... 113
HR4-900 ............................................... 122
HR4-902 ............................................... 121
HR4-904 ......................... 118-119, 121-122
HR4-911 .................................................122
HR4-913 ............................................... 122
HR4-914 ............................................... 122
HR4-915 ............................................... 126
HR4-917 ............................................... 126
HR4-919 ............................................... 126
HR4-921 ............................................... 126
HR4-923 ............................................... 126
HR4-925 ............................................... 126
HR4-928 ............................................... 126
HR4-929 ............................................... 126
HR4-931 ............................................... 126
HR4-933 ............................................... 126
HR4-935 ............................................... 126
HR4-937 ............................................... 126
HR4-939 ............................................... 126
HR4-941 ............................................... 126
HR4-943 ............................................... 153
HR4-951 ............................................... 130
HR4-953 ............................................... 124
HR4-955 ............................................... 124
HR4-957 ............................................... 124
HR4-959 ............................................... 124
HR4-961 ............................................... 124
HR4-963 ............................................... 124
HR4-965 ............................................... 124
HR4-969 ............................................... 123
HR4-971 ............................................... 123
HR4-973 ............................................... 123
HR4-975 ............................................... 123
HR4-977 ............................................... 123
HR4-979 ............................................... 123
HR4-981 ............................................... 127
HR4-983 ............................................... 127
HR4-985 ............................................... 127
HR4-987 ............................................... 127
HR4-989 ............................................... 127
HR4-991 ............................................... 127
HR4-993 ............................................... 124
HR4-995 ............................................... 124
HR4-997 ............................................... 124
HR4-999 ............................................... 124
H R 4 - C O N T I N U E D
HR5-102 ............................................... 134
HR5-104 ............................................... 129
HR5-106 ............................................... 129
HR5-108 ............................................... 129
HR5-110 ............................................... 129
HR5-112 ....................................... 120, 129
HR5-113 ............................................... 129
HR5-114 ............................................... 129
HR5-118 ............................................... 129
HR5-120 ............................................... 129
HR5-210 ............................................... 168
HR5-211 ............................................... 167
HR5-212 ............................................... 168
HR5-213 ............................................... 167
HR5-214 ............................................... 172
HR5-215 ............................................... 166
HR5-219 ............................................... 170
HR5-221 ............................................... 172
HR5-223 ............................................... 168
HR5-224 ............................................... 166
HR5-230 ............................................... 167
HR5-231 ............................................... 173
HR5-233 ............................................... 169
HR5-237 ............................................... 170
HR5-239 ............................................... 172
HR5-241 ............................................... 170
HR5-900 ............................................... 125
HR5-902 ............................................... 125
HR6-100 ............................................... 151
HR6-102 ............................................... 151
HR6-104 ............................................... 139
HR6-106 ............................................... 139
HR6-108 ............................................... 139
HR6-110 ............................................... 139
HR6-112 ............................................... 139
HR6-114 ............................................... 139
HR6-116 ............................................... 139
HR6-118 ............................................... 139
HR6-120 ............................................... 139
HR6-122 ............................................... 139
HR6-124 ............................................... 139
HR6-126 ............................................... 139
HR6-128 ............................................... 139
HR6-130 ............................................... 139
HR6-132 ............................................... 139
HR6-134 ............................................... 139
HR6-136 ............................................... 139
HR6-138 ............................................... 139
HR6-140 ............................................... 139
HR6-142 ............................................... 139
HR6-144 ............................................... 139
HR6-146 ............................................... 139
HR6-148 ............................................... 139
HR6-150 ............................................... 139
HR6-151 ............................................... 139
HR6-152 ............................................... 139
HR6-154 ............................................... 139
HR6-156 ............................................... 139
HR6-158 ............................................... 139
HR6-160 ............................................... 139
HR6-162 ............................................... 139
HR6-164 ............................................... 139
HR6-166 ............................................... 139
HR6-168 ............................................... 139
HR6-170 ............................................... 139
HR6-172 ............................................... 139
HR6-174 ............................................... 139
HR6-175 ............................................... 139
HR6-177 ............................................... 139
HR6-179 ............................................... 139
HR7-099 ............................................... 179
HR7-100 ............................................... 176
HR7-102 ............................................... 176
HR7-104 ............................................... 178
HR7-106 ............................................... 178
HR7-108 ............................................... 177
HR7-110 ............................................... 177
HR8-002 ............................................... 123
HR8-004 ............................................... 123
HR8-006 ............................................... 123
HR8-008 ............................................... 123
HR8-010 ............................................... 123
HR8-012 ............................................... 123
HR8-014 ............................................... 123
HR8-028 ....................................... 145, 152
HR8-030 ............................................... 140
HR8-032 ............................................... 140
HR8-056 ................................................. 90
HR8-058 ................................................. 90
HR8-061 ............................................... 171
HR8-062 ............................................... 169
HR8-069 ............................................... 101
HR8-072 ............................................... 125
HR8-074 ................................................. 93
HR8-076 ................................................. 93
HR8-078 ................................................. 93
HR8-080 ................................................. 93
HR8-082 ................................................. 93
HR8-084 ................................................. 93
HR8-086 ............................................... 169
HR8-088 ................................................. 91
HR8-090 ................................................. 91
HR8-092 ......................................... 57, 104
HR8-094 ............................................... 120
HR8-098 ................................................. 94
HR8-100 ............................................... 171
HR8-102 ............................................... 125
HR8-104 ............................................... 125
HR8-106 ............................................... 125
HR8-108 ............................................... 125
HR8-110 ............................................... 125
HR8-112 ............................................... 120
HR8-114 ............................................... 120
HR8-116 ............................................... 120
HR8-118 ............................................... 120
HR8-120 ............................................... 120
HR8-122 ............................................... 120
HR8-124 ............................................... 120
HR8-126 ............................................... 120
HR8-128 ............................................... 120
HR8-130 ............................................... 120
HR8-132 ............................................... 120
HR8-133 ............................................... 115
HR8-134 ................................................. 82
HR8-135 ................................................. 82
HR8-136 ................................................. 82
HR8-137 ................................................. 82
HR8-138 ................................................. 82
HR8-139 ................................................. 82
HR8-140 ................................................. 82
HR8-141 ................................................. 82
HR8-146 ................................................. 82
HR8-147 ................................................. 82
HR8-148 ................................................. 83
HR8-149 ................................................. 83
HR8-150 ................................................. 78
HR8-152 ................................................. 78
HR8-154 ................................................. 78
HR8-156 ................................................. 78
HR8-162 ................................................. 92
HR8-163 ................................................. 92
HR8-164 ................................................. 92
HR8-165 ................................................. 92
HR8-166 ................................................. 92
HR8-167 ................................................. 92
HR8-168 ................................................. 92
HR8-169 ................................................. 92
HR8-170 ................................................. 92
HR8-171 ................................................. 84
HR8-172 ................................................. 84
HR8-173 ............................................... 118
HR8-174 ............................................... 118
HR8-175 ............................................... 118
HR8-176 ............................................... 118
HR8-177 ............................................... 118
HR8-178 ............................................... 118
HR8-179 ............................................... 118
HR8-180 ............................................... 118
HR8-181 ............................................... 118
HR8-182 ............................................... 119
HR8-184 ............................................... 119
HR8-186 ............................................... 119
HR8-188 ............................................... 119
HR8-190 ............................................... 119
HR8-192 ............................................... 119
HR8-194 ............................................... 119
HR8-196 ............................................... 119
HR8-198 ............................................... 119
DI-038 ..................................................... 89
DI-039 ..................................................... 89
DI-040 ................................................... ..89
DI-041 ..................................................... 89
DI-043 ..................................................... 80
H R 6 -
H R 7 -
H R 8 -
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Hampton Research Phone: 800-452-3899 or 949-425-1321 Fax: 949-425-1611 www. hampt onresearch. com Cust omer Servi ce: i nf o@hrmai l . com Techni cal Support : t ech@hrmai l . com
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MSDS
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