Periodic Acid-Schiff (PAS) Reaction
Technique: Cut paraffin sections at 6 microns.
Dissolve 1 g of basic fuchsin in 200 cc hot distilled water.
Bring to boiling point. Cool and add 2 g potassium
metabisulfite, and 10 cc normal hydrochloric acid. Let
bleach for 24 hours, then add 0.5 g activated carbon.
Shake 1 minute and filter through coarse filter paper.
Repeat filtration until solution is colorless. Store in a
Schiff’s Leuko-Fuchsin Solution
Dissolve 1 g basic fuchsin in 200 cc hot distilled water. Bring
to boiling point. Cool to 50oC. Filter and add 20 cc normal
hydrochloric acid. Cool further and add 1 g anhydrous
sodium bisulfite, or sodium metabisulphite. Keep in the
dark for 48 hours until solution becomes straw colored.
Test for Schiff’s Leuko-Fuchsin Solution
Pour a few drops of Schiff’s solution into 10 cc of 37–40%
formaldehyde in a watch glass. If the solution turns
reddish purple rapidly, it is good. If the reaction is delayed
and the resultant color deep blue-purple, the solution is
Normal Hydrochloric Acid Solution
Hydrochloric acid, Concentrated specific gravity
0.2% Light Green counterstain (Stock)
Light green, stock solution 10 cc
Distilled water, sterile 100 cc
Store in refrigerator. Good for 1 week.
(Instead of diastase solution, one can use one’s own saliva).
(If PAS reaction with digestion is needed, place
sections in 0.5% diastase for 20 minutes. Saliva can
also be used. Rinse in running tap water 10 minutes,
5. Periodic acid solution for 5 minutes (oxidizer).
7. Place in Coleman’s Feulgen or Schiff’s leuko-fuchsin
8. Place in running tap water for 10 minutes for pink
9. Stain in Harris’s hematoxylin for 6 minutes or light
green counterstain for a few seconds. Light green is
recommended for counterstaining sections in which
fungi are to be demonstrated. Omit steps 10 through
11. Differentiate in acid alcohol—3 to 10 quick dips.
13. Dip in ammonia water to blue sections.
14. Wash in running tap water for 10 minutes.
16. Absolute alcohol—2 changes.
Glycogen, mucin, hyaluronic acid, reticulin, fibrin or
thrombi, colloid droplets, hyaline of arteriosclerosis,
preserved, most basement membranes, colloid of pituitary
stalks and thyroid, amyloid infiltration and other elements
may show a positive reaction—rose to purplish red.
Background—pale green (with light green counterstain).
Technique: Cut paraffin section at 6 microns.
Hydrochloric acid, concentrated 1 cc
Equal parts of solutions A and B. Prepare fresh.
Aluminum chloride, anhydrous 0.5 g
Mix stain in small test tube. Heat over small flame for 2
minutes. Liquid becomes almost black and syrupy. Dilute
with 100 cc of 50 % alcohol and let stand for 25 hours. Filter.
Dilute 1 to 4 with tap water for use.
1. Deparaffinize sections through 2 changes of xylene,
absolute and 95% alcohol to distilled water as usual.
Remove mercury precipitates through iodine and
2. Stain for 7 minutes in working solution of Weigert’s
3. Wash in tap water for 5 to 10 minutes.
4. Place in diluted mucicarmine solution for 30 to 60
Check control slide with microscope after 30 minutes.
5. Rinse quickly in distilled water.
6. Stain in metanil yellow solution for 1 minute.
7. Rinse quickly in distilled water.
8. Rinse quickly in 95% alcohol.
9. Dehydrate in 2 changes of absolute alcohol, clear with
2 to 3 changes of xylene, and mount in DPX.
Technique: Cut paraffin sections at 6 microns.
Crystal violet-to saturateapproximately 14 g
Working Crystal Violet Solution
Crystal violet, stock solution 10 cc
Hydrochloric acid, concentrated 1 cc
Dissolve with the aid of heat.
If this medium cannot be obtained, then put a drop of water
on the section to be seen and examine under microscope.
1. Deparaffinize sections through 2 changes of xylene,
absolute and 95% alcohol to distilled water as usual.
2. Stain in working crystal violet solution for 1 to 2
minutes. Use control slide. Check with microscope.
5. Seal edges of coverslip with nail polish.
Von Kossa’s Method for Demonstrating Calcium
Fixation: Alcohol or 10% formalin. Alcohols are preferred.
Technique: Cut paraffin sections at 6 microns.
Solution stable only for 2 weeks.
5% Sodium Thiosulfate Solution
Nuclear Fast Red (Kernechtrot) Stain
Dissolve 0.1 g nuclear fast red powder in 100 cc of a 5%
aqueous aluminum sulfate solution with the aid of heat.
Cool and filter. Add a crystal of thymol as preservative.
Keeps well at room temperature. Can be reused.
1. Deparaffinize sections through 2 changes of xylene,
absolute and 95% alcohols to distilled water.
2. Place in 5% silver nitrate solution for 30 to 60 minutes
exposed to direct sunlight, ultraviolet lamp, or a 100
watt desk lamp light. Use chemically clean container.
4. Place in 5% solution thiosulfate for 2 to 3 minutes.
5. Wash well in distilled water.
6. Counterstain in nuclear fast red for 5 minutes.
8. Dehydrate with 2 changes of 95% alcohol, absolute
alcohol, clear with 2 changes of xylene and mount in
Ziehl-Neelsen Stain for Acid-Fast Bacteria
Technique: Paraffin sections at 4 to 6 microns.
Store at room temperature. Filter before use.
Hydrochloric acid, concentrated 1 cc
Sulfuric acid concentrated 1 cc
Working Methylene Blue Solution
Glacial acetic acid, concentrated 0.5 cc
1. Deparaffinize sections through 2 changes of xylene,
and run through absolute and 95% alcohols to distilled
water as usual. Remove mercury precipitates through
iodine and hyposolutions, if necessary.
2. Stain sections with carbol fuchsin solution for 10
minutes. Filter solution before use.
4. Decolorize with 1% acid alcohol or 1% sulfuric acid
water, until sections are pale pink.
5. Wash thoroughly with running water for 8 minutes.
6. Counterstain by dipping one slide at a time in working
methylene blue solution. Sections should be pale blue.
Overstaining will mask bacilli.
7. Wash with tap water and distilled water.
8. Dehydrate with 2 changes of 95% alcohol and absolute
alcohol, clear with 2 changes of xylene, and mount in
Erythrocytes—yellowish-orange.
Other tissue elements—pale blue.
Modular enclosed tissue processing system:
¾ A unique, safe, totally enclosed system that will not
release vapors into the laboratory during processing
¾ Keyboard programing and memory with video display
and simple touch button controls accept ordinary
¾ Separate command, reaction and storage modules
offer flexible choice of layouts
¾ One command module can control up to 5 reaction and
storage modules independently and simultaneously
¾ Nine programs per reaction module—total capacity 45
programs, each of which can be recalled instantly
¾ Programs can be monitored at every stage.
806 Concise Book of Medical Laboratory Technology: Methods and Interpretations Histocenter (Shandon)
Integrated tissue embedding center:
¾ Provides efficient utilization of bench space—easy to
clean work surfaces all on one level
¾ Incorporates processed tissue storage, cassette opening
¾ Wax temperature can be preset and monitored
continually on digital display—can also be switched to
¾ Wax dispenser can be foot operated leaving both hands
¾ Wax flow rate easily adjusted with nozzle heated to
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