 Don’t use these reagents if the label is not available or damaged.

 Do not use dark glass slide.

 Don’t use the kit if damaged or the glass vials are broken or leaking
and discard the contents immediately.
 Test materials and samples should be discarded properly in a
biohazard container.
Blood Grouping Reagents:  Wash hands and the test table top with water and soap once the
Anti-A Monoclonal Reagent, Anti-B Monoclonal testing is done.
Reagent, Anti-AB Monoclonal Reagent, Anti-D  Heamolysed blood sample should not be used for testing.
 The test should be performed at room temperature in a well let
IgG/IgM blend Reagent, & Their variants area with very good visibility.
SLIDE AND TUBE TESTS  Failure to follow the procedure in this package insert may give false
results or safety hazard.
For In-Vitro and professional use only  Close the vial tightly after each test.
 The reagent is considered toxic, so don’t drink or eat beside it.
Store at 2- 8°C  If spillage of reagent occurs clean with disinfectant (disinfectant
INTENDED USE used could be irritable so handle with care).
The blood grouping reagents are used to detect the presence or absence
of A, B or Rhesus Antigens on the surface of human red blood cells based STORAGE CONDITIONS
on hemaglutination using slide or tube test techniques in whole blood  The reagents should be stored refrigerated between 2 - 8°C.
samples or anticoagulant blood samples collected in EDTA , citrate or  Never Freeze or expose to elevated temperature.
heparin tubes.  The reagent is stable until the expiry date stated on the product
label. Do not use the reagents past the expiry date.
INTRODUCTION & PRINCIPLES
Blood grouping reagents are prepared from In-Vitro culture supernatants REAGENT PREPRATION
of hybridized immunoglobulin-secreting mouse cell lines. The reagents are  The reagents are intended for use as supplied, no prior preparation
diluted with phosphate buffer containing sodium chloride, EDTA and or dilution of the reagent is required.
bovine albumin to give reagents that are optimized for use in tube and  All reagents should be brought to room temperature before use.
slide procedures. Anti-A monoclonal reagent is colored with acid blue
(patent blue) dye, Anti-B monoclonal reagent is colored with acid yellow SPECIMEN COLLECTION AND PREPARATION
(tartrazine) dye, and Anti-AB monoclonal reagent is not colored. The test  Blood collected with or without anticoagulant (EDTA, Heparin or
procedure is based on hemaglutination principle, where red cells Citrate) can be used for Antigen typing.
possessing the antigen agglutinate in the presence of the corresponding Note: Blood collected without anticoagulant should be tested
antibody indicating that the result is positive. The test is considered immediately.
negative when no agglutination appears.  The specimens should be tested as soon as possible after collection.
Anti-D IgG/IgM blend reagent is prepared from carefully blended human If testing is delayed, the specimens should be stored at 2- 8 ºC,
monoclonal IgM and IgG. Anti-D IgG/IgM blend reagent is suitable for slide Sample must be retained to room temperature prior to analysis.
and tube test procedures. The reagent will directly agglutinate Rh D (Testing should be carried out within five days of collections).
positive cells, including majority of variants (but not D VI) and a high  Insure that there is no sign of hemolysis.
proportion of weak D (Du) phenotypes. The reagent will agglutinate  At the time of the test, centrifuge the blood sample at 1200 RCF for
category DVI and low grade weak D (Du) phenotypes by the indirect 3 minutes.
anti-globulin techniques.  Blood collection is to be done with great care.
Anti-D IgG/IgM blend reagent is diluted with a sodium chloride solution,
sodium phosphate solution and bovine albumin (sodium caprylate free). PROCEDURES
Anti-D IgG/IgM blend reagent is not colored. The procedure is based on A. DIRECT TUBE METHOD AT ROOM TEMPERATURE
hemaglutination principle, where red cells’ possessing the antigen 1. Prepare a 5% suspension of red blood cells in isotonic solution.
agglutinates in the presence of the corresponding antibody in the reagent 2. Using the vial dropper, transfer a drop (40±10µl) of each reagent
indicating that the result is positive. The test is considered negative when into a separate and appropriately marked tube.
no agglutination appears. 3. Add 50 µl of red blood cell suspension prepared in step 1.
4. Shake to homogenize the mixture, then centrifuge at 500g for 1
MATERIALS minute.
MATERIALS PROVIDED 5. Gently shake the tube in such a way to detach the cell pellet and
Blood Grouping Reagents: macroscopically observe for any possible agglutination.
 Anti-A monoclonal reagent (10 ml/vial), Clone: (9113D10). 6. Read the reaction immediately.
 Anti-B monoclonal reagent (10 ml/vial), Clone: (9621A8). 7. For Anti-D tube, if the reaction is weak or negative, shake the tubes
 Anti-AB monoclonal reagent (10ml/vial), Clone: and incubate at 37°C for 15 minutes.
(152D12+9113D10). 8. Wash the red blood cells twice with isotonic saline solution (NaCl
 Anti-D IgG/IgM Blend reagent (10 ml/vial), Clone: (P3X61 + 0.9%) and discard the last washing liquid.
P3X21223B10 + P3X290 + P3X35). 9. Add one drop (50µl) of the AHG reagent into the tube. Mix and
MATERIALS NEEDED BUT NOT PROVIDED centrifuge at 120g for 1 minute.
 Plastic test tube. 10. Gently shake the tube in such a way to detach the cell pellet and
 Isotonic buffered saline (pH 6.9). macroscopically observe for any possible agglutination.
 Applicator sticks. 11. Read the reaction immediately.
 Centrifuge (100-1200 RCF for tube test). B. ANTIGLOBULIN INDIRECT METHOD for ANTI-D
 Timer. 1. After immediately centrifuging and reading as above, if the reaction
 Incubator is weak or negative, shake the tubes and incubate at 37°C for 15
 Anti-Human Globulin Reagent (can be ordered from Atlas Medical). minutes.
 White or transparent glass slide. 2. Wash the red blood cells twice with isotonic saline solution and
discard the last washing liquid.
PRECAUTIONS 3. Add one drop (40 µl ± 10 µl) of ANTI-HUMAN GLOBULIN to the
 The reagents are intended for in vitro diagnostic use only. tube. Mix and centrifuge at 120 RCF for 1 minute.
 The test is for well trained professional healthy user not for lay 4. Gently shake the tube in such a way to detach the cell pellet and
user. macroscopically observe for any possible agglutination.
 These reagents are derived from animal and human sources, thus, 5. Read the reaction immediately.
appropriate care must be taken in the use and disposal of these C. DIRECT SLIDE METHOD AT ROOM TEMPERATURE
reagents, as there are no known test methods that can guarantee 1. Bring reagents and samples to room temperature (18-25°C).
absence of infectious agents. 2. Using the wax pen divide the slide into appropriate numbers of
 Do not use reagents if it is turbid or contain particles as this may divisions.
indicate reagent deterioration or contamination. 3. Using the provided dropper, place one drop (40 µl ± 10 µl) of each
 Protective clothing should be worn when handling the reagents. reagent onto its correspondent division on the slide.
 The reagents contain (0.1-0.2%) Sodium Azide and 0.02% sodium 4. Add 25µl of the precipitated cells next to each drop of reagents .
arseniate which is toxic and can be absorbed through the skin. 5. Mix the reagent and the cells using a clean stirring stick over an
When drained, the drains should be thoroughly flushed with area with a diameter of approximately 20-40mm.
water. 6. Incubate the slide at room temperature (18-25°C) without stirring
 The reagents should be used as supplied and in accordance to the for 30 seconds.
procedure mentioned below. Don’t use beyond expiration date. 7. Hold the slide and gently rock the slide for 3 minutes and observe
 Avoid cross contamination of reagents or specimens. macroscopically for any agglutination.
 Visible signs of microbial growth in any reagent may indicate 8. Read the reaction immediately.
degradation and the use of such reagent should be discontinued.
 READING THE RESULT

Compliance
Lot A

Lot B
POSITIVE: If Agglutination appears.

Lot C
CE marked
NEGATIVE: If no agglutination is observed. device
Use the below table to determine the blood group:

Result of each reaction 61 61 61 61 100%

Tube Technique
monoclonal

monoclonal

monoclonal

IgG/IgM
ABO

Anti-AB
reagent

reagent

reagent

reagent
Anti-D
Anti-A

Anti-B

blend
Group Group B
Positive with anti-B monoclonal reagent and anti-AB
monoclonal reagent
+ - + + A+ Negative with anti-A and Negative control
+ - + - A-

Compliance
Lot A

Lot B

Lot C
- + + + B+ CE marked
- + + - B- device
+ + + + AB+
+ + + - AB- 61 61 61 61 100%
- - - + O+
- - - - O-
Slide Technique
Group O
STABILITY OF THE REACTIONS
 ABO Blood Grouping Tube tests should be read immediately Negative with anti-A monoclonal reagent, Anti-B
following centrifugation. monoclonal reagent and anti-AB monoclonal reagent
 Slide tests should be interpreted within three minutes to avoid the Negative with Negative control
possibility that a negative result may be incorrectly interpreted as

Compliance
Lot A

Lot B

Lot C
positive due to drying of reagents. CE marked
 Delay in reading and interpreting results may result in weekly device
positive or falsely negative reactions. Slide tests should be
interpreted at the end of the three minutes. 241 241 241 241 100%

PROCEDURE LIMITATION Tube Technique

1. False positive/ negative results may occur due to: Group O
 Contamination from test materials.
Negative with anti-A monoclonal reagent, Anti-B
 Improper storage, cells concentration, incubation time or
monoclonal reagent and anti-AB monoclonal reagent
temperature.
Negative with Negative control
 Improper or excessive centrifugation.

Compliance
 Deviation from the recommended technique. Lot A

Lot B

Lot C
CE marked
 Blood samples of weak A or B subgroups may give rise to false
device
negative results or weak reactions.
2. Weaker reactions may be observed with stored blood than with
fresh blood. 243 243 243 243 100%
3. ABO antigens are not fully developed at birth, weaker reactions
may therefore occur with cord or neonatal red cells. Slide Technique
4. ABO blood grouping interpretation on individuals greater than 6
Group AB
months old should be confirmed by testing serum or plasma of the
Positive with anti-A monoclonal reagent, Anti-B
individual against group A and group B red cells (reverse grouping).
monoclonal reagent and anti-AB monoclonal reagent
If the results obtained with the serum do not correlate with the red
Negative with Negative control
cell test, further investigation is required. Compliance
5. Return the kit to the agent if it does not function properly.
Lot A

Lot B

Lot C

6. Anti-D IgG/IgM blend Reagent tests conducted on particular CE marked

weak-D phenotypes, while satisfactory, cannot ensure recognition device
of all weak variants, due to the variability of antigen patterns.
33 33 33 33 100%
DIAGNOSTIC PERFORMANCE CHARACTERISTICS Tube Technique
The following tables compare the results in slide and tube techniques of 3
Group AB
lots of Atlas Medical reagents and the results of a CE marked device.
Positive with anti-A monoclonal reagent, Anti-B
Slide Technique
monoclonal reagent and anti-AB monoclonal reagent
Group A Negative with Negative control
Positive with anti-A monoclonal reagent and anti-AB
Compliance

monoclonal reagent
Lot A

Lot B

Lot C

CE marked
Negative with anti-B and Negative control device
Compliance
Lot A

Lot B

Lot C

CE marked
24 24 24 24 100%
device
No inversion in diagnosis has been shown: from a qualitative point of view
we have observed 100% compliance in direct group testing in slide and
232 232 232 232 100% tube techniques for determination of A, B, AB and O groups for the three
Tube Technique lots of Atlas Medical.
Group A
QUALITY CONTROL
Positive with anti-A monoclonal reagent and anti-AB
The reactivity of all blood grouping reagents should be confirmed by
monoclonal reagent
testing known positive and negative red blood cells on each day of use.
Negative with anti-B and Negative control
To confirm the specificity and sensitivity, Blood grouping reagents should
Compliance

be tested with antigen-positive and antigen-negative red blood cells.

Lot A

Lot B

Lot C

CE marked
device REFERENCES
1. BCSH Blood Transfusion Task Force. Guidlines for microplate
212 212 212 212 100% techniques in liquid-phase blood grouping and antibody screening.
Clin. Lab. Haem 1990: 12, 437-460.
2. Issitt P. D. Applied Blood Group Serology, 3rd ed. Miami: Montgomery
Slide Technique
Scientific, 1985.
Group B 3. Kholer G., Milstein C. Continuous culture of fused cells secreting
Positive with anti-B monoclonal reagent and anti-AB antibody of predefined specificity, 256, 495-497, 1975
monoclonal reagent 4. Messeter L. et. al. Mouse monoclonal antibodies with anti-A, anti-B
Negative with anti-A and Negative control and anti-A,B specificities, some superior to human polyclonal ABO
reagents, Vox Sang 46, 185-194, 1984
5. Race R.R. and Sanger R. Blood groups in man, 6th ed., Oxford:
Blackwell Scientific, 1975.
6. Voak D. ET. al., Monoclonal anti-A and anti-B development as cost
 effective reagents. Med. Lab. Sci 39, 109-122. 1982. Ludwig-Erhard-Ring 3
7. Standards for Blood Banks d Transfusion Service. 11th Ed., 15827 Blankenfelde-Mahlow
Washington D.C., AABB 1984:25. Germany
8. Widmann F.K.ed Technical Manual, 9th Ed., Wahington D.C.: AABB Tel: +49 - 33708 – 3550 30
1985:9. Email: Info@atlas-medical.com
PPI861A01
Rev J (03.09.2019)
LIST OF VARIENTS:
Product Code Product Name
8.02.00.0.0010 Anti-A Monoclonal Reagent (Titer: 1 /512), 10ml/vial, 1 vial/Carton Box
8.02.00.1.0100 Anti-A Monoclonal Reagent (Titer: 1 /512), 10ml/vial. 10 vials / Plastic Pack
8.02.00.1.0180 Anti-A Monoclonal Reagent (Titer: 1 /512), 10ml/vial. 18 vials / Carton Box
8.02.01.0.0010 Anti-B Monoclonal Reagent (Titer: 1 /512), 10ml/vial, / Carton Box
8.02.01.1.0100 Anti-B Monoclonal Reagent (Titer: 1 /512), 10ml/vial, 10 vials / Plastic Pack
8.02.01.1.0180 Anti-B Monoclonal Reagent (Titer: 1 /512), 10ml/vial, 18 vials / Carton Box
8.02.02.0.0010 Anti-AB Monoclonal Reagent (Titer: 1 /512), 10ml/vial, 1 vial/ Carton Box
8.02.02.1.0100 Anti-AB Monoclonal Reagent (Titer: 1 /512), 10ml/vial, 10 vials/Plastic Pack
8.02.02.1.0180 Anti-AB Monoclonal Reagent (Titer: 1 /512), 10ml/vial, 18 vials/Carton Box
8.02.03.0.0010 Anti-D IgG/IgM Blend Reagent (Titer: 1 /128), 10ml/vial, 1 vial/ Carton Box
8.02.03.1.0100 Anti-D IgG/IgM Blend Reagent (Titer: 1 /128), 10ml/vial, 10 vials / Plastic Pack
8.02.03.1.0180 Anti-D IgG/IgM Blend Reagent (Titer: 1 /128), 10ml/vial, 18 vials / Carton Box
8.02.04.0.0010 Anti-A Monoclonal Reagent (Titer: 1 /256), 10ml/vial, 1 Vial/Carton Box
8.02.04.0.0100 Anti-A Monoclonal Reagent (Titer: 1 /256), 10ml/vial, 10 vials / Plastic Pack
8.02.05.0.0010 Anti-B Monoclonal Reagent (Titer: 1 /256), 10ml/vial, 1vial/Carton Box
8.02.05.0.0100 Anti-B Monoclonal Reagent (Titer: 1 /256), 10ml/vial, 10 vials /Plastic Pack
8.02.05.6.0030 ABO Set (Anti-A (1/256), Anti-B (1 /256), Anti-D (1/64)),3x10ml / plastic Pack
8.02.05.7.0020 ABO Set: Anti-A (1/256), Anti-B (1 /256), 2x10ml /Plastic Pack
8.02.06.0.0010 Anti-AB Monoclonal Reagent (Titer: 1 /256), 10ml/vial, 1vial/Carton Box
8.02.06.1.0100 Anti-AB Monoclonal Reagent (Titer: 1 /256), 10ml/vial,10 vials /Plastic Pack
8.02.06.1.0180 Anti-AB Monoclonal Reagent (Titer: 1 /256), 10ml/vial,18 vials / Carton Box
8.02.07.0.0010 Anti-D IgG/IgM Blend Reagent (Titer: 1 /64), 10ml/vial, 1Vial/ Carton Box
8.02.07.1.0100 Anti-D IgG/IgM Blend Reagent (Titer: 1 /64), 10ml/vial, 10 vials / Plastic Pack
8.02.47.0.0030 ABO Set (Anti-A (1 /512), Anti-B (1 /512), Anti-D (1 /128)),3x10ml/Plastic Pack
8.02.47.1.0030 ABO Set (Anti-A (1 /256), Anti-B (1 /256), Anti-D (1 /64)), 3x10ml /Carton Box.
8.02.47.3.0030 ABO Set (Anti-A (1 /256), Anti-B (1 /256), Anti-D (1 /64)), 3x10ml /Plastic Pack
8.02.47.5.0030 ABO Set (Anti-A (1 /256), Anti-B (1 /256), Anti-D (1 /128)), 3x10ml/Plastic Pack
8.02.49.0.0040 ABO Set (Anti-A (1 /256), Anti-B (1 /256), Anti-AB (1 /256), Anti-D (1 /64)), 4x10ml/Carton Box
8.02.49.2.0040 ABO Set (Anti-A (1 /256), Anti-B (1 /256), Anti-AB (1 /256), Anti-D (1 /128)), 4 x 10ml, 4 vials/Plastic Pack
8.02.53.0.0040 ABO Set (Anti-A (1 /512), Anti-B (1 /512), Anti-AB (1 /512) Anti-D (1 /128)), 4x10ml/Plastic Pack
8.02.53.1.0040 ABO Set (Anti-A (1 /512), Anti-B (1 /512), Anti-AB (1 /512) Anti-D (1 /128)), 4x10ml, 4vials/Plastic Pack
8.02.70.0.0010 Anti-A monoclonal reagent , Titer (1/1024), 10 ml/vial, 1Vial/ Carton Box
8.02.71.0.0010 Anti-B Monoclonal reagent (Titer: 1 /1024) , 10 ml/vial ,1Vial/ Carton Box
8.02.72.0.0010 Anti-AB Monoclonal reagent (Titer: 1 /1024) , 10 ml/vial , 1Vial/ Carton Box
8.02.85.0.0010 Anti-D IgG/IgM Blend reagent ( Titer 1 /256), 10ml/vial, 1Vial/ Carton Box

Catalogue Number Temperature limit

In Vitro diagnostic medical
Caution
device
Contains sufficient for <n> Consult instructions
tests and Relative size for use (IFU)
Batch code Manufacturer

Fragile, handle with care Use-by date

Do not use if
Manufacturer fax number
package is damaged
Manufacturer
telephone number Date of Manufacture

Keep away from sunlight Keep dry