HISTOPATH MTAP 2 (Meeting 1) 11.

Frozen section:
- (-)10 to (-)20 degrees Celsius
PRE-ANALYTICAL PREPARATION - Commonly used in:
1. Tissue degradation starts as soon as its blood supply is interrupted. (TRUE) o Rapid pathologic diagnosis during surgery
- Warm (initial response) and cold (in preservative) ischemia o Diagnostic and research enzyme histochemistry
- Small (no need to resection) and large (need to dissect for better penetration) o Diagnostic and research demonstration of soluble substances such as lipids,
specimens carbohydrates
o Immunofluorescent and immunohistochemical staining
2. Warm ischemia is beyond the control of the histopathology laboratory. (TRUE) o Some specialized silver stains, particularly in neuropathology
- Other factors such as speed and skills of surgeon (taken in consideration especially in
frozen section) 12. Most scrupulous attention to the accurate and unambiguous identification of sample
pathology data base (FALSE)
3. Prevention of cold ischemia requires immediate transport of tissue from the operating room
to the laboratory (TRUE) - Number of specimen
- FS, Imprints, IHC (Immunochemistry), lymphoma, mol and cytogenetic studies - Barcoding
- Slide or smear preparations are for screening, used to identify for integrity of sample - Multiple ways of ID
- Pull apart and smears are used to determine the delay in transport - ID of same specimen –DNA ID
- Most errors are from clerical error
4. The proper ration of fixative to tissue is 5:1. (FALSE)
- No amount of troubleshooting can solve poor fixation 13. Barcoding specimens greatly helps in specimen identification along with patient
- Proper ratio is 10:1 or 20:1 demographics (TRUE)
- Immediately place to fixative (30 mins to 1 hr) - Minimizes errors in Clerical and typographical errors

5. Poorly fixed tissue will not process, cut or stain well. (TRUE) 14. False negative results may happen when the tissue used for targeting therapy testing is not
- Most especially in case of Ischemia and autolysis properly fixed (TRUE)
- Integrity of sample is not good
6. ASCO – CAP guidelines recommend a fixation time of a minimum 6 hrs and maximum of 48 - Caused by decomposition leading to misdiagnosis
hrs (TRUE)
- Thickness of 3-5 mm 15. Rapid fixing of tissue
- Large specimen sectioned 5 mm interval - Liquid nitrogen = cracks due to rapid expansion of the ice within tissue, producing ice
- Hollow organ should be opened crystals or freeze artifacts. It also overcools urgent biopsy blocks causing damage to both
- For Breast cancer block and blade if sectioning is done at (-)70 C or below
- Vapor phase to form around the tissue , acting as an insulator that causes uneven
7. A properly filled-out histopathology request form includes patient demographics, history, cooling of tissue, particularly of muscle biopsies and making diagnostic interpretation
pertinent physical and laboratory as well as imaging findings to help pathologists arrive at difficult. This problem can be overcome by freezing the tissues in ISOPENTANE, OCT OR
the correct tissue diagnosis (TRUE) FREON 2.2 that has high thermal conductivity
- Do not practice Gamesmanship (superiority or advantage from other doctors) - Cold Knife Procedure = uses CO2 cylinder
- Cryostat Procedure – Mounting of Tissue Block = synthetic water-soluble glycols and
8. Weight of intact specimen is rounded to the nearest (0.1 g) resins are generally used as mounting media for tissue blocks that need to be sectioned
- Thickness is rounded to the nearest 0.1 cm in cryostat. The OCT (optimal cutting temperature) compound is used in CRYOSTAT
- NON FATTY BREAST TISSUE (-15 TO -25= TEMP)
9. Used to identify and orient the specimen’s components (Incking) - FATTY BREAST TISSUE (-35 = TEMP)
- Red, yellow, orange inks are common - Routine processing: Paraffin
- Sutures = for long lateral and short superior - Freezing Previously Fixed Tissue – clean slides should be coated with albumin or
- Nicking = for lateral orientation chrome-glycerin jelly so that the fixed tissue will attach to the slide (SUSCEPTIBLE IN
WASHING OUT)
10. Request is completed by consultant - Examination of nerve and muscle – separate portions that will allow formalin fixation
- (INFORMATION NEEDED BY THE PATHOLOGIST) Pre-operative, intra-operative finding, and paraffin embedding, unfixed snap-frozen for cryostat sections, fixation and resin
specimen submitted, matter obtained, procedure and post operative and imaging used embedding for electron microscopy (fixed in buffered solution of glutaraldehyde and
- History of patient, and initial diagnosis postfixed in osmium tetroxide)
- If chemical fixation of tissue blocks is to be avoided, namely:
 o Freeze-drying-rapid freezing (quenching) of fresh tissue at (-)160 C and FIXATION
subsequently removing ice water molecules (dessication) by transferring the still 1. Dehydrant coagulant fixatives that has a closed structure with water but way more
frozen tissue block into a vacuum chamber at a higher temperature e.g. (-)40C dangerous? (METHANOL)
(sublimation) without the use of any chemical fixative a. Alcohol has closed structure with water
o QUENCHING, DESSICATION, SUBLIMATION b. Methanol can cause blindness and kidney disease
o Freeze substitution instead of being subjected to dehydration in an expensive
vacuum drying apparatus, is fixed in Rossman’s formula or in 1% Acetone and 2. A formalin fixed tissue sample shows yellowish pigment upon microscopic examination. To
dehydrated in absolute alcohol prevent occurrence of such deposit one should: (Filter the formalin before use)
a. Review solution that removes pigmentation
SPECIMEN PREPARATION i. Lillie’s method
1. For lymph node, specimens that require flow cytometric methods or immunophenotyping, ii. Picric Acid with formalin
where should the specimen be placed? (TRANSPORT MEDICINE) iii. Kardasewitsch’s method
a. LYMPH NODE- Most important in planning therapeutic prognosis iv. 1.5% Iodine = for mercury pigmentation
b. Needs to be Fresh.
c. Bivalved = formalin (if processed immediately) 3. For electron microscopy, tissue should be fixed for _3_ hours and placed in holding buffer
d. Sentinel Lymph Node = first to involved in metastasis, usually in breast cancer (3)
e. Large should be dissected = 1cm interval (bread loafing), ensures area are identified. a. Only small samples
b. 0.2 or 0.3 samples placed in glutaraldehyde.
2. The following are used for the gross examination of specimens except: (FILTER PAPER) we
use LENS PAPER (to avoid washing-out) 4. Made up of only one component substance (SIMPLE FIXATIVE)
a. 1st step = specimen ID / same name accession number
b. Small malignant specimen = wrong container so clean area and cutting tools to avoid 5. Formalin diffuses into the tissue at the rate of __ mm per hour (1 mm)
contamination a. Prevent inaccurate diagnosis, maintain cell relationship, stroma (collagen, reticulum,
c. Weight = 0.1 g (hyperplastic thymic tissue) elastin, amorphous substances)
d. Dimension= 0.1 cm b. Refractive index (transparency of tissue) and hardening (advantage in fixation for
e. Color, consistencity easy handling of sample)
f. Small fragment-inked, uses lens paper for cell block, and to cover it then put it in c. Carbohydrates and lipids in fixative disintegrates or shrinks, have particular fixatives
cassesttes so it wont wash out
6. Recommended mainly for tumor biopsies especially for the skin (HEIDENHAIN’S SUSA
3. Which of the following is not true of specimen sectioning? (ALL LYMPH NODE SPECIMEN SOLUTION)
SHOULD BE FIXED IN NEUTRAL BUFFERED FORMALIN) because it should be FRESH a. Orth’s Fluid = study of early degenerative processes and tissue necrosis
a. LN = submitted fresh as much as possible b. Potassium Dichromate = preserves lipids and mitochondria
b. With tumor = Id the following c. Chromic Acid = precipitate all CHON and fixes carbohydrates
c. Site the size, location, and structure invaded, vascular imvasion, distance from d. Regaud’s (Moller) Fluid = for demonstration of Golgi bodies
resection margin, LN
d. Cassette: 3x2.5x4 cm 7. Fixative –additive (add themselves in tissue conformation of protein or macromolecules),
e. Not >.3 cm (NOT BULKY), with paper tag non-additive (without combining organic compound such acetone and alcohols which
f. Colon; proper orientation dissociating water)
a. Coagulant (create network, penetrate the interior of the tissue, zinc salt, mercuric
4. Which of the following is true of loading specimens in cassettes? (Specimens must not be chloride, picric, ethyl, methyl and acetone), non-coagulant (create gel that difficult
more than 0.3 cm) to penetrate by subsequent solutions; needs to cut thinly)
a. Secondary identifier inside, don’t use sign pen. Use pencil b. Factors contribute to fixation
i. Temp: room temperature to 45C (increase temp, increase activity)
5. Grosses criteria of specimen rejection ii. Non coag + large specimen = needed to be cut thinly
a. Request and specimen label discrepancy iii. Size: 3 mm sectioning
b. No label or mislabel iv. Volume: 15-20
c. Leaking v. Time: within 20-30 mins and not more than 60 mins
d. No history or clinical data vi. Fixation time: 6 to 48 hrs in 10% NBF
e. Inappropriate ID of specimen vii. ASCP-CAP FOR human Epidermal Receptor-2 (found in breast, increase
indicates IHC)
8. Recommended for demonstration of chromatin, mitochondria, mitotic figures, Golgi c. Spray cans of alcohol fixatives are marketed to physicians doing PAP smears.
bodies, RBCs and colloid-containing bodies (REGAUD’s FLUID) d. Ethanol = most unstable DNA fragments for polymerase chain reaction,
a. 10% NBF formaldehyde limits the size of fragments which can be retrieved
i. Fixative of choice, IHC (detects ANTIBODY) and FISH e. Mercurials fix tissues through an unknown mechanism that increases staining
ii. Preparation: formalin fastest penetrating capacity, 3.6/hr, 7%2/4 hrs brightness and gives excellent nuclear detail. However, it penetrates poorly (long
(temp:24, pH 7); cross-linking is complete in 48 hrs fixing time) and produce tissue shrinkage. Their best application is for fixation of
iii. Osmolality: Hyper = shrink, hypo = swell, iso = ill effect hematopoietic and reticuloendothelial tissues (metallic staining)
iv. Used of gauze with saline and ice for transport f. Oxidizing agent = permanganate fixatives which stabilize tissue structure with
v. Nucleus: RNA= 45 deg ; DNA = 65 deg extensive effects like denaturation and structure; uses secondary fixative
vi. Nuclear bubbling = formalin effect in nucleus formation of chromatin strand g. Chromate fixative = decomposition of fixative, penetrates all tissue, preserve
during deparaffinization step carbohydrates
vii. Protein = configuration of bonds h. Picric fixative = for connective tissue, glycerin, extract lipid for superior salt in
viii. Lipid = osmium tetroxide and chromic acid immunostaining
i. Acetic acid = precipitate DNA, for nuclear protein
9. Which of the following is incorrect regarding fixation of tissue? (LARGER TISSUE REQUIRE j. Lead fixative = more on enzyme activity
LESS FIXATIVE AND SHORTER FIXATION TIME)

10. Commonly used for bone marrow biopsies (B-5 FIXATIVE) DECALCIFICATION
a. Consider the stain used 1. Notes
b. Fix in alcohol if smear a. Calcium ions
c. If bone, need to decalcify b. Surface decalcification
i. Releasing is 6-8 days c. Ideal cut for hard specimens (bone and myoma) 2-3 mm for hard tissue
ii. First 3 days for decalcification step d. Bone marrow = buffered formalin (alternative: zinc formalin mixtures, b-5, formol-
iii. Picric or formalin +buffer acetic alcohol [Davidson’s] Or Bouin’s solution
e. Strong Mineral Acids = hydrochloric (for surface) or nitric acid at concentrations up
11. Mercuric deposits may be removed by immersing tissues in alcoholic iodine prior to to 10% with acids to form soluble calcium salts
staining, through a process known as de-zenkerization f. Weaker organic acids = lactic, acetic
g. Chelating agent = EDTA which combines with calcium, forming an insoluble non-
12. Notes ionized complex for 1-3 weeks for small specimens but may take 6-8 weeks (not
a. Secondary fixation is the process of placing an already fixed tissue in a second used na since it takes time)
fixative. May be done before dehydration and deparaffinized sections before h. Bone marrow biopsy = core (decalcified) and smear (stained with Wright Giemsa)
staining. Usually with 10% formalin or 10% formol saline as primary fixative i. Ion exchange resin – removing calcium ions from formic acid containing decalcifying
b. Post chromatization is a form of secondary fixation (potassium dichromate) solution. Not recommended for fluids containing mineral acids such as nitric acid or
whereby a primarily fixed tissue is placed in aqueous solution of 2.5 to 3% potassium hydrochloric acid
dichromate for 24 hrs to act as mordant i. 20-30 times the volume (large amount of resin)
c. Washing out is the process of removing excess fixative from the tissue after ii. Stays in solution for 1-14 days
fixation in order to improve staining and remove artifacts from the tissues (Kelly’s, iii. Removes ions from acid
Zenker’s, and Flemming’s solutions, formalin, osmic acid and 50-70% alcohol and j. Electrophoresis (electrical ionization)
alcohol iodine) i. Positively charged calcium ions are attracted to negative electrode and
d. Crush artifacts may be found in surgical specimens particularly in liver biopsies, subsequently removed from decalcifying solution shortened due to heat
associated with an intense eosinophilic staining at the center of the tissue in H&E and electrolytic reaction (same with chelating but with electricity)
stained sections. This may be due to artificial coagulation of partially fixed protein by k. Tissue softeners = Perenyi’s fluid may act as both decalcifying agent and tissue
ethanol or by incomplete wax impregnation during subsequent histological softener.
processing i. Molliflex
e. Heat effect = zonal fixation effect; large sample is not cut so inner part will not be ii. 2% hydrochloric acid
fixed. Difference morphological and staining characteristic between inside and iii. 1% hydrochloric acid in 70% alcohol
outside.
2. Recommended for routine decalcification of post mortem research tissue (FORMIC ACID)
13. Protein denaturants (methanol, ethanol, acetone) a. Usually 100%
a. Rarely used alone for fixing blocks unless studying nucleic acids b. Best all around decalcifier
b. Good for cytologic smears because they act quickly and give good nuclear details c. Incorporate in formalin and buffer
 iv. Cellosolve – rapid combustible at 110-120F are toxic
3. Bone marrow continues to undergo mitosis up to (30 mins) mins after death when v. Triethyl phosphate – as dehydrating solution in the staining sequence
refrigerated. vi. Tetrahydrofuran—dehydrates and clears
vii. For EM – ethanol as dehydrating solvent and propylene as transition fluid
4. Inadequate decalcification may result in the poor cutting of hard tissues and damage to the
knife-edge during sectioning (TRUE) CLEAR
a. Honing and stropping 1. Notes
a. De-alcoholization
5. EDTA (versene) is the commonly used chelating agent (TRUE) b. Optical clarity or transparency to the tissue due to their relatively high refractive
index
6. Used as surface decalcification or remedy for grating sensation when sectioned with a c. In frozen sections, glycerin and gum syrum are used when tissue is to be cleared
microtome knife (10% HYDROCHLORIC ACID) direct,y from water
d. Hydrocarbons – flammables, solution with most organic solvent, paraffix wax,
7. Calcium may be removed by the ff agents except: (ALCOHOLIC ACID) coagulates nitrocellulose

8. Formol Nitric Acid imparts yellow color that will impair the staining reaction of the cells and 2. What are the effects of incomplete clearing? (UNEVEN H&E STAINING AND POOR
can be prevented by adding (5% SODIUM SULFATE AND 0.1% UREA) CHROMATIN PATTERNS (patches; pale or intense))

9. Recommended for teeth and small pieces of bone (VON EBNER’S FLUID) 3. Embedding media will not infiltrate tissue that contains water (TRUE)

10. Optimum temperature for decalcification (18-30 C) 4. Incomplete dehydration effect/s (SOFT, MUSHY AND CLEARING AGENT NOT ACT
PROEPRRLY DUE TO WATER) ALL OF THE CHOICES

11. Dehydration should evaporate very fast (FALSE) 5. The tissue has an average refractive index of ___ to __ while most synthetic resins have a
refractive index ranging from 1.51 to 1.55 (1.53 to 1.54)
12. Primarily employed for blood and tissue films and for smear preparation (METHYL
ALCOHOL) 6. As the refractive index of mounting medium that approaches that of the tissue, the tissue
becomes more and more transparent (TRUE)
13. What method used when tissue is wrapped in a gauze bag and suspended in a bottle
containing dioxane and a little anhydrous calcium oxide (WEISEBERGER’S METHOD) 7. In frozen sections, tissue is to be cleared directly from water (no de-alcholozation) process
involved (TRUE)
- GRAUPNER METHOD (dioxane only)
8. All of the following are characteristics of good clearing agent, except (should dissolve out
DEHYDRATE aniline dyes) SHOULD NOT
1. Notes
a. Isopropyl (IPA) – versatile substitute, efficient lipid solvent, soluble with water, 9. Clearing time for xylene (30 MINS TO 1 HR; 2 changes of xylene so max of 2 HRS)
organic solvent and melted paraffin
b. Glycol = ethers 10. Xylene becomes milky when an incompletely dehydrated tissue is immersed in it (TRUE)
c. Ethoxyethanol, cellosolve (phototensi), poluethylene glycols (dehydrate and embed)
d. Acetone (4degrees Celsius) rapid and coag secondary fix, best for fatty specimen, 11. Used as a substitute for xylene or benzene for clearing both during embedding and
raw material for shabu mounting processes (TOLUENE)
e. Tetrahydrofuran = highly volatile and flammable, best universal solvent and less
toxic than dioxane 12. Most carcinogenic or may damage bone marrow (BENZENE)
f. Dioxane = toxic and carcinogenic formed peroxide may explode upon air exposure
g. Commonly used Dehydrating Agents Are 13. All of the following makes tissue transparent, except (CHLOROFORM)
i. Alcohol – most common and different decal process according to the type of
fixative used 14. Recommended for tough tissues (CHLOROFORM)
ii. Acetone – not recommended for routine dehydration process (shrinkage in
2 hrs max) 15. Recommended for clearing embryos, insects and very delicate specimen (ANILINE OIL)
iii. Dioxane – ribbon poorly, extremely dangerous, uses Graupner’s and
Weiseberger’s method
 16. Notes
a. Xylene (xylol) – less than 5 mm, toxic and narcotic
b. Toluene – clearing 1-2 hours, expensive Microtomy
c. Benzene – it’s use for clearing purposes is therefore strongly discouraged 15. Used to reduce surface tension and allow the section to flatten out (WATER BATH; fishing
d. Chloroform – up to 1 cm, thickness – skin, fibroid and decalcified tissues for nervous out)
tissues, lymph nodes and embryos 16. Primarily used for cutting tissue sections at __ micra for electron microscopy (0.5)
e. Cedarwood Oil – recommended for central nervous system tissues and cytological 17. Invented by Minot (ROTARY MICROTOME)
studies, particularly of smooth muscle and skin, x2 changes every 2-3 days 18. Recommended for frozen sections or for cutting extremely hard and tough specimen (PLANE
f. Aniline Oil – recommended for clearing embryos, insects and very delicate WEDGE KNIFE)
specimens 19. Designed for ultrathin section microtome, the specimen is examined under an EM (GLASS OR
g. Clove Oil – slow and difficult GEM)
h. Carbon Tetrachloride – similar to that of chloroform (cheaper) 20. The perfect optimum cutting angle is obtained when the sides of the wedge knife are
i. Tetrahydrofuran – two processes at the same time, processing time and allowing inclined at an angle of about (15)
more time for fixation, dehydrates and clears 21. Mineral oil is not recommended and should never come in contact with a strop (TRUE)
j. Dioxane – miscible both with water and paraffin, toxic to human especially to the 22. Rocking microtome that is resting on pivots and a supporting column and attached to
liver micrometer screw (lower arm – nylon, it is moving)
k. Terpenes are isoprene polymers found in essential oils originally derived from 23. Best manual sharpening (Belgium yellow)
plants, though some are now synthesized – substitute only 24. To prevent uneven sections or alternate thin and thick sections the knife should be inclined
with a __ clearance angle from cutting plane (5-10)
Impregnation and Embedding 25. Line up the tissue in the proper position to the knife (Ratchet – mechanical part)
1. Holds the cell and intercellular structures in their proper relationship while thin sections are 26. Toe to heel direction, edge last (Stropping – polishing)
cut (INFILTRATION) 27. Notes
2. Embedding also known as (BLOCKING, CASTING, ORIENTATION) a. Rocking Microtome – for cutting serial sections pf large blocks of paraffin
3. In automatic processing wax bath thermostats should be set at least __ degrees above the embedded tissues
melting point of the wax (3) b. Rotary Microtome – for cutting paraffin embedded sections
4. In automatic processing only 2-3 changes of wax are required to remove the clearing and c. Sliding Microtome – for cutting celloidin embedded sections
poorly impregnate the specimen, why (DUE TO CONSTANT AGITATION) d. Freezing microtome – for cutting unembedded frozen sections
5. Used for routine work temperature of paraffin wax (56C, deteriorate if above 65) e. Cryostat or cold microtome – for cutting frozen sections
6. The degree pf the vacuum should not exceed __ mmHg (500 mmHg() f. Ultrathin microtome – for cutting sections for EM
7. If laboratory temperature is from 20-24C paraffin-wax with a melting point of (54-58C)
8. Paraffin oven must be maintained at temperature 2 to 5C above melting point of paraffin to Cutting Sections
be used for impregnation (TRUE) 1. Notes
9. Consists of two L-shaped strips of heavy brass or metal arranged on a flat metal plate a. Paraffin – may be cut by rocking and rotary microtome
(LEUCKHARTS) b. Celloidin – cut by means of sliding microtome
10. Larger and denser tissue blocks usually require longer periods and more frequent changes c. Frozen – from fixed and frozen with CO2 or for fresh or fixed tissues frozen with
of wax (TRUE) cryostat
11. Vacuum chamber is usually maintained at a temperature of __ above the melting point of d. Trimming
the wax (2-4C) e. Faults and Problems Observed during Section Cutting
12. Difficult to remove clearing medium in tissue (CHLOROFORM AND OTHER OILS) 2. This dehydrating agent is used to remove residual ethanol in the preparation of tissue
13. If lab temperature is 15-18C – the melting point of wax should be between __ (50 – 54C) samples for EM (PROPYLENE OXIDE)
14. Notes 3. Used as the primary fixative in TEM (2.5% GLUTARALDEHYDE)
a. Impregnation and embedding medium namely 4. A type of stain I EM wherein the electron beam primarily interacts with the stain, when the
i. Paraffin wax – 39 to 68C improve by above 60 to 65. 55 to 58C – Paraplast stain is added to sample, the stain surrounds the sample which then appear on a dark
and embeddol background (NEGATIVE STAIN)
ii. Celloidin (colloidin) – suitable for specimens with large hollow cavities which 5. Electron microscopes can achieve a resolution up to (2 MILLION)
tend to collapse, for hard and dense tissues such as bones and teeth and for 6. This type of fixative reacts with unsaturated lipids and stabilizes tissue proteins during post
large tissue sections of the whole embryo—days or weeks fixation (1% OSMIUM TETROXIDE)
iii. Gelatin – when dehydration is to be avoided and when tissues are to be 7. In processing tissue EM, this procedure is done to prevent the tissue sample from
subjected to histochemical and enzyme studies putrefaction and deterioration (FIXATION)
iv. Plastic – renal biopsies and bone marrow biopsies, epoxy, polyester or 8. A type of EM that shows a 3D images of a sample (STEM)
acrylic 9. Produces a largely magnified image by using electrons instead of light to form image (EM)
 10. Sectioning of tissue blocks for EM is done with the use of glass or diamond knife in a
(ULTRAMICROTOME) Rapid diagnosis
11. NOTES 1. For early treatment, convenient practice
a. Electron beams are used in EM to illuminate the specimen and thus create and 2. Notes:
image wavelength of electrons are 100 000 times shorter than visible lights 3. Microwave antigen retrieval that has a higher temperature processing means (SHORTER
b. Parts of TEM HEATING TIME TO ACHIEVE STRONG INTENSITY OF STAIN)
i. Electron source 4. Microwave antigen retrieval is used to unmask or retrieve antigens using 10mm citrate
ii. Electromagnetic lens system buffer at ph 8.0. Microwave reduced significantly the time required for staining of tissue,
iii. Sample holder particularly when using heavy metals. (BOTH TRUE)
iv. Imaging system 5. Composed of methanol and polyethylene glycol? (UMFIX)—universal
6. Haste processing of tissue, except (CHANGING OF USED REAGENT)—done every week
7. Stimulates alcohol diffusion in histological processing (
Adhesives and Mounting Media 8. Processor allows addition of new specimen every 15 mins as a reaction chamber becomes
1. No more adhesives since use of oven and xylene available (CONTINUOUS TISSUE PROCESSOR)
2. Possible that sections may float from the slide 9. Optimum temperature used in metallic staining during rapid tissue processing (75 to 95C)
3. Mounting media used for preparation that has been dehydrated and cleared in xylene or 10. Fixative that can be used in freeze-substitution (1%ACETONE AND ROSSMAN FLUID)
toluene and are recommended for majority of staining methods (RESINOUS) 11. Paraffin must be added to the microwave in liquid form in a CRTP (TRUE)
4. Importance except (DETERIORATION OF XYLENE) 12. One major disadvantage of RTP techniqueis the shortened time intervals in between each
5. Active coloring agent I hematoxylin (HEMATIN) step that causes uneven processing of the tissue sample (FALSE)
6. Protects the stained section from getting scratched and from bleaching pr deterioration due 13. In rapid processing technique, lower temperature allows shorter heating time to achieve
to oxidation (MOUNTING MEDIUM) strong intensity of staining (False)
7. Chief solvent used for stain except (HEMATOXYLIN) 14. Rapid tissue processor reduces processing time to 3 hours when combined with microwave
8. Notes fixation (TRUE)
a. Commonly used adhesive: ALBUMIN + THYMOL 15. Microwave processing allows the use of alcohol during dehydration using ine step only
b. Albumin = retains stain and gives dirty background eliminating the used of icreasingalcohol volume (TRUE)
c. Poly-l-lysine = 0.1% solution and further diluted (1:10 with dH2O) 16. Notes:
d. Amino propyltriethoxy s (APES) is a better section adhesives and coated slides can a. Paraffin must be added to microwave in luquid form
be stored for a longtime b. Melted with intermedium at 82C for ethyl alcohol, and 0C for isopropanol
e. Aqueous Mounting Media c. Purpose: FLASH EVAPORATION of alcohol or propanol. Alcohol quickly heats and
i. Water dissipates while the paraffin remains inert, allowing paraffin to fully infiltrate the
ii. Glycerin –preservative specimen, eliminating the use of xylene
iii. Farrant’s – Gum Arabic BONE MARROW AND CYTOLOGY
iv. Apathy’s – gum Arabic 17. Bone marrow smear preparation can used (BOTH SQUASH AND SPREAD)
v. Bruin’s – glucose 18. Histotechnologist are authorized to extract bine marrow from patients (FALSE)
f. Resinous Mounting Media 19. Ferric iron merges with potassium ferricyanide, giving a blue stained reaction commonly
i. Canada Balsam = for whole mounts and thick sections (does not shrink known as (PRUSSIAN BLUE)
much) 20. Other stain used to demonstrate plasma cells (METHYL GREEN PYRONIN STAIN)
ii. DPX = dibutyl phthalate and xylene 21. Counterstain of perl’s prussian blue (NUCLEAR FAST RED)
iii. XAM 22. For inclusion bodies (GIEMSA STAIN)
23. No specimen is aspirated (APLASTIC ANEMIA)
Principles of Staining 24. Esophageal washing is (NON-GYN WASHING)
1. Notes 25. Notes
a. Silver Nitrate = for impregnation and staining agent a. Indication of Bone Marrow Biopsy
b. Intravital Staining = injecting the dye i. No material obtained by aspiration (dry tap)
c. Counterstaining = different color of stain to provide contrast background ii. Bone marrow structure visualization: lipid storage disease, malignancy,
d. Metachromatic = specific dyes which differentiate particular substances by staini g metastatic disease
them in a color different from the stain b. BM preparation: touch and smear prep
e. Regressive = staining of H&E i. Air dried
f. Progressive = diffused color and obscured details type of staining ii. Fixed
g. Indirect Staining = used link or bridge between the tissue and dye iii. Stained by Wright’s or Giemsa
h. Direct = example: methylene and eosin
 iv. Other stain: Methyl Green pyronin = differentiation of DNA and RNA.
Reticulin stain= reticulum fiber
c. Optimal cell preservation for cytology studies = 95% ethanol or equal parts of 95%
ethanol and ether
d. If smears from effusions cannot be made immediately, 50% alcohol and carbowax
e. If fluid specimen is enough for cytocentrifugation, prepare at least 2 cytocentrifuged
smears and a cell block
f. PAP smear
i. Membrane filtration = Thin Prep technique
ii. Density gradient centrifugation and gravity sedimentation = SurePath
technique
iii. Conventional Spatula = 360 degree roll in cervix