Stability of Special Staining Solutions

STABILITY OF SPECIAL STAINING SOLUTIONS

General Guidelines Concerning Solutions

1. Check stock solutions regularly for signs of deterioration.

2. All solutions should be dated and initialed by the person who prepared the solution.

3. Use caution to prevent cross solution contamination.

4. Use distilled or double distilled water to prepare solution.

5. Use reagent grade chemicals.

6. Use dyes which are certified by the Biological Stain Commission.

7. Keep solutions in well stoppered bottles at all times.

8. Refrigerated solutions should be poured into the staining jar, and the bottled solution returned to the refrigerator before it warms to room temperature.

9. Return all staining solutions to shelves soon after use to prevent mix-up.

10. Always use clean glassware.

11. Make sure that balance, pH meter, etc., are in proper working order.

12. The following chemicals should be kept refrigerated:

1) All buffer solutions

2) Hydroquinone

3) Paraldehyde keeps well in the freezing compartment. It will freeze to a solid, but can be easily melted at room temperature.

4) Phosphomolybdic acid

5) Phosphotungstic acid

6) Protargol

7) Silver nitrate and all solutions containing silver nitrate

The following information indicates the stability of the staining solutions used in most of the staining procedures in this manual. The shelf-life indicated for the various solutions is based upon our many years of experience with the staining methods in which these solutions are used. There may be some variation of the shelf-life indicated because of the temperature in the room or refrigerator in which they are stored and because of the quality of the distilled water, chemicals and the dyes with which they have been prepared. Some of the solutions may be used several weeks or months longer than what is indicated.

An asterisk preceding the solution indicates it is used for one day and then discarded.

Ziehl-Neelsen (AFB)

1. Carbol fuchsin -------- 6 months

2. *Acid alcohol -------- 6 months

3. Methylene blue -------- 6 months

Fite (AFB)

1. Mineral oil - xylene -------- 1 year

2. Carbol fuchsin -------- 6 months

3. *Acid alcohol-------- 6 months

4. Methylene blue -------- 6 months

Truant (AFB)

1. Auramine - rhodamine -------- 6 months

2. *Acid alcohol -------- 6 months

3. Eriochrome black T -------- 6 months

Brown-Brenn (bacteria)

1. Hucker-Conn crystal violet -------- 1 year

2. *Grams iodine -------- 2 months

3. *Ethyl alcohol - acetone-------- 6 months

4. *Basic fuchsin -------- 3 months

5. *Picric acid - acetone -------- 6 months

6. *Acetone - xylene -------- 6 months

Gram (bacteria)

1. Hucker-Conn crystal violet -------- 1 year

2. *Grams iodine -------- 2 months

3. *Safranin O, stock and working solution -------- 1 year

Wolbachs Giemsa

1. Acid alcohol -------- 6 months

2. Giemsa, stock -------- 1 year

3. 10% Triton X-100 -------- 6 months

4. *Giemsa, working

5. 1% Acetic acid-------- 6 months

Gomori-Grocott (GMS)

1. Chromic acid -------- 1 year

2. Sodium borate -------- 6 months

3. Methenamine - silver nitrate, stock (refrigerate) -------- 1 month

4. *Methenamine - silver nitrate, working

5. 10% Triton X-100-------- 6 months

6. *Sodium bisulfite -------- 2 months

7. Gold chloride -------- 6 months

Keep gold chloride in an amber bottle. Gold chloride in a Coplin jar should be kept in a dark place until ready for use because it is sensitive to light.

8. Sodium thiosulfate -------- 6 months

9. Fast green, stock -------- 6 months

10. Fast green, working -------- 2 weeks

Churukians Ammoniacal Silver (fungi and pneumocystis carinii)

1. Periodic acid -------- 6 months

2. Ammoniacal silver (refrigerate) -------- 1 month

3. Gold chloride -------- 6 months

4. Sodium thiosulfite -------- 6 months

5. Fast green, stock -------- 6 months

6. Fast green, working -------- 2 weeks

Warthin-Starry (bacteria)

1. Acetic acid-glycine, stock (refrigerate) -------- 6 months

2. *Acetic acid-glycine, working

3. *Silver nitrate

4. *Gelatin

5. *Hydroquinone

6. *Silver nitrate - gelatin - hydroquinone

7. Tartarazine-------- 6 months

Steiner (spirochetes)

1. Uranyl nitrate -------- 2 months

2. *Silver nitrate

3. Gum mastic -------- 6 months

4. *Hydroquinone

5. *Reducing solution

Alcian Blue (acid mucopolysaccharides)

1. Acetic acid -------- 6 months

2. Alcian blue -------- 6 months

3. Nuclear fast red -------- 4 months

Alcian Blue - PAS (acid and neutral mucopolysaccharides)

1. Acetic acid -------- 6 months

2. Alcian blue -------- 6 months

3. Periodic acid -------- 6 months

4. Modified Lillies Schiff solution (refrigerate) -------- 6 months

Kreyberg (acid mucopolysaccharides)

1. Alcian blue -------- 6 months

2. Ammonia - alcohol -------- 2 months

3. Weigerts hematoxylin, stock -------- 6 months

4. Weigerts ferric chloride - hydrochloric acid -------- 6 months

5. Weigerts working -------- 1 week

6. Acid alcohol -------- 6 months

7. Erythrosin B -------- 2 months

8. Metanil yellow -------- 6 months

PAS and PAS with Diastase (glycogen)

1. Periodic acid -------- 6 months

2. Modified Lillies Schiff solution (refrigerate)-------- 6 months

3. *Diastase

4. Lillie-Mayer hematoxylin -------- 1 year

Southgates Mucicarmine (acid mucopolysaccharides)

1. Weigerts hematoxylin, stock -------- 6 months

2. Weigerts ferric chloride - hydrochloric acid -------- 6 months

3. Weigerts working -------- 1 week

4. Mucicarmine, stock (refrigerate)-------- 4 months

5. *Mucicarmine, working

6. Tartarazine -------- 6 months

Periodic Acid - Thionin - KOH - PAS (PAT)

1. *Periodic acid

2. *Thionin

3. *Potassium hydroxide

4. Modified Lillies Schiff solution (refrigerate) -------- 6 months

Best Carmine (glycogen)

1. Weigerts hematoxylin, stock -------- 6 months

2. Weigerts ferric chloride - hydrochloric acid -------- 6 months

3. Weigerts working -------- 1 week

4. Carmine, stock (refrigerate) -------- 4 months

5. *Carmine, working solution -------- 3 days

6. *Differentiating solution

Puchtlers Congo Red (amyloid)

1. Weigerts hematoxylin, stock -------- 6 months

2. Weigerts ferric chloride - hydrochloric acid -------- 6 months

3. Weigerts working-------- 1 week

4. 1% Sodium hydroxide -------- 1 year

5. Congo Red -------- 2 months

Lieb Crystal Violet (amyloid)

1. Crystal violet, stock -------- 1 year

2. Crystal violet, working -------- 6 months

3. Apathys mounting media -------- 10 years

Jones (basement membranes)

1. Periodic acid -------- 6 months

2. Methenamine - silver nitrate, stock (refrigerate) -------- 1 month

3. Sodium borate -------- 6 months

4. *Methenamine - silver nitrate, working

5. Gold chloride (keep in the dark) -------- 6 months

6. Sodium thiosulfate -------- 6 months

7. Nuclear fast red -------- 4 months

Alcian Blue - PAS (kidney)

1. Acetic acid -------- 6 months

2. Alcian blue -------- 6 months

3. Periodic acid -------- 6 months

4. Modified Lillies Schiff solution -------- 6 months

Gomori Trichrome

1. Bouins -------- 2 months

2. Weigerts hematoxylin, stock -------- 6 months

3. Weigerts ferric chloride - hydrochloric acid -------- 6 months

4. Weigerts working -------- 1 week

5. Trichrome -------- 4 months

6. Acetic acid -------- 6 months

Masson Trichrome

1. Bouins -------- 2 months

2. Weigerts hematoxylin, stock -------- 6 months

3. Weigerts ferric chloride - hydrochloric acid -------- 6 months

4. Weigerts working -------- 1 week

5. Biebrich scarlet - acid fuchsin -------- 2 months

6. Phosphomolybdic - phosphotungstic acid -------- 6 months

7. Aniline blue -------- 2 months

8. Acetic acid -------- 6 months

Modified Massons (Cardiac Tissues)

1. Alcoholic picric acid -------- 6 months

2. Ponceau S -------- 6 months

3. Phosphomolybdic acid -------- 6 months

4. Aniline blue -------- 2 months

5. Acetic acid -------- 6 months

Mallory (PTAH)

1. *Potassium permanganate -------- 1 week

2. *Oxalic acid -------- 6 months

3. Zenkers, stock -------- 6 months

4. Zenkers, with acetic acid -------- 2 weeks

5. Lugols iodine -------- 6 months

6. *Sodium thiosulfate -------- 6 months

7. Phosphotungstic acid hematoxylin (PTAH) -------- 1 year

The PTAH solution is generally considered to have a long shelf-life. However,

the quality of the staining results should be carefully checked for any signs of deterioration.

Van Gieson (collagen)

1. Weigerts hematoxylin, stock -------- 6 months

2. Weigerts ferric chloride - hydrochloric acid -------- 6 months

3. Weigerts working -------- 1 week

4. Acid fuchsin -------- 6 months

5. Picric acid -------- 6 months

Verhoeff (elastic)

1. Hematoxylin, 5% alcoholic -------- 1 year

2. Ferric chloride, 10% -------- 6 months

3. Lugols iodine -------- 6 months

4. *Verhoeffs, working

5. Tartarazine-citric acid -------- 6 months

6. Acid fuchsin -------- 6 months

7. Picric acid -------- 6 months

Frazer-Lendrum (fibrin)

1. Weigerts iron hematoxylin, working -------- 1 week

2. Orange G - Picric acid-------- 1 year

3. Acid fuchsin -------- 6 months

4. Tartarazine, differentiating -------- 1 year

5. Light green -------- 1 year

Carstairs (fibrin and platelets)

1. Weigerts iron hematoxylin, working -------- 1 week

2. Picro-orange G -------- 1 year

3. Ponceau-fuchsin -------- 6 months

4. Aniline blue -------- 1 year

Gomori (reticulum)

1. *Acidified potassium permanganate -------- 3 days

2. *Potassium metabisulfite -------- 2 months

3. Ammoniacal silver (refrigerate) -------- 7 days

4. *Formalin -------- 6 months

5. Gold chloride (keep in the dark) -------- 6 months

6. Sodium thiosulfate -------- 6 months

7. Nuclear fast red -------- 4 months

Microwave Ammoniacal Silver (argentaffin, melanin)

1. *Ammoniacal silver -------- 1 month

2. Gold chloride (keep in the dark) -------- 6 months

3. Sodium thiosulfate -------- 6 months

4. Nuclear fast red -------- 4 months

Schmorl (reducing substance)

1. *Ferric chloride

2. *Potassium ferricyanide

3. Nuclear fast red -------- 4 months

Churukian-Schenk (argyrophil cells)

1. Citric acid - Sodium citrate, stock -------- 1 year

2. *Citric acid-sodium citrate, working

3. *Silver nitrate

4. *Bodians developer

5. Nuclear fast red -------- 4 months

Giemsa (plastic bone marrow and smears)

1. Hydrochloric acid-alcohol -------- 6 months

2. Triton X-100 -------- 6 months

3. Phosphate buffer, pH 6.6 (refrigerate) -------- 1 year

4. Giemsa, stock -------- 1 year

5. *Giemsa, working

Orcein (HBsAg)

1. *Acidified potassium permanganate

2. *Oxalic acid -------- 6 months

3. Orcein -------- 6 months

Aldehyde Fuchsin (HBsAg)

1. *Acidified potassium permanganate

2. *Oxalic acid -------- 6 months

3. Aldehyde fuchsin -------- 2 months

Toluidine Blue (mast cells)

1. *Potassium permanganate -------- 1 month

2. Potassium metabisulfite -------- 2 months

3. Toluidine blue -------- 6 months

Dahal (calcium)

1. *Alizarin red S -------- 1 year

2. Fast green, stock -------- 6 months

3. Fast green, working -------- 2 weeks

Rhodanine (copper)

1. *Rhodanine, stock

2. *Rhodanine, working

3. Lillie-Mayer hematoxylin -------- 1 year

4. *Sodium borate -------- 6 months

Pearl (iron)

1. Hydrochloric acid -------- 3 months

2. Potassium ferrocyanide -------- 3 months

3. Triton X-100-------- 6 months

4. *Hydrochloric acid - potassium ferrocyanide

5. Nuclear fast red -------- 4 months

Feulgen (DNA)

1. 5N hydrochloric acid -------- 1 year

2. Modified Lillies Schiff solution (refrigerate) -------- 6 months

3. *Fast green

Methyl Green - Pyronin (DNA-RNA)

1. Buffer solutions -------- 1 year

2. Methyl green -------- 6 months

3. Pyronin Y -------- 6 months

Aldehyde Fuchsin (pancreatic beta cells)

1. Lugols iodine -------- 6 months

2. Sodium thiosulfate -------- 6 months

3. Aldehyde fuchsin -------- 2 months

4. Fast green, stock -------- 2 months

5. Fast green, working -------- 2 weeks

Wilson-Ezrin (pituitary)

1. Periodic acid-------- 2 months

2. Modified Lillies Schiff solution (refrigerate) -------- 6 months

3. Weigerts hematoxylin, stock -------- 6 months

4. Weigerts ferric chloride - hydrochloric acid -------- 6 months

5. Weigerts working -------- 1 week

6. Acid alcohol -------- 6 months

7. Orange G -------- 6 months

8. *Phosphotungstic acid -------- 6 months

Hall (bilirubin)

1. Trichloroacetic acid -------- 6 months

2. Ferric chloride -------- 6 months

3. *Fouchets

4. Acid fuchsin -------- 6 months

5. Picric acid -------- 6 months

Lison (hemoglobin)

1. Patent Blue V (stock ) -------- 1 week

2. *Patent Blue V (working)

3. Nuclear fast red -------- 4 months

Lillies Nile Blue (melanin and lipofuscin)

1. Nile blue -------- 6 months

Melanin Bleach

1. *Potassium permanganate -------- 1 week

2. Oxalic acid -------- 6 months

3. Nuclear fast red -------- 4 months

Gomori (urates)

1. Methenamine - silver nitrate, stock -------- 1 month

2. Boric acid -------- 6 months

3. Sodium borate -------- 6 months

4. *Methenamine - silver nitrate - buffer

5. Gold chloride (keep in the dark) -------- 6 months

6. Sodium thiosulfate -------- 6 months

7. Fast green, stock -------- 6 months

8. Fast green, working -------- 2 weeks

Bodian (nerve fibers)

1. *Protargol-------- 48 hours

2. *Reducing

3. Gold chloride (keep in the dark) -------- 6 months

4. *Oxalic acid -------- 6 months

5. Sodium thiosulfate-------- 6 months

Cresyl Violet Acetate (nissl substance)

1. Acetic acid -------- 6 months

2. *Cresyl violet acetate

Cresyl Violet (helicobacter pylori)

1. Cresyl violet ------------------------------------------------------------------ 3 months

2. Phosphoric acid -------------------------------------------------------------- 6 months

Einarson (nissl substance)

1. Gallocyanin -------- 1 week

Lapham (myelin and glial fibers)

1. Gallocyanin -------- 1 week

2. Phloxine -------- 1 month

3. Fast green-------- 2 months

4. Acetic acid-------- 6 months

Luxol Fast Blue - Cresyl Violet acetate (myelin and nerve cells)

1. Luxol fast blue -------- 1 year

2. Cresyl violet acetate, stock -------- 1 month

3. *Cresyl violet acetate, working

4. Lithium carbonate -------- 1 year

Thioflavin S (amyloid in neurofibrillary plaques) and Thioflavin T

1. Lillie-Mayer hematoxylin -------- 1 year

2. *Thioflavin S -------- 1 week

3. Thioflavin T ----------------------------------------------------------------- 6 months

4. Acetic acid -------- 6 months

Naphthol AS-D Chloroacetate (leukocyte esterase)

1. Pararosanilin, hexazotized (refrigerate) -------- 1 year

2. Sodium nitrite (refrigerate)-------- 1 year

3. Buffer solutions (refrigerate) -------- 1 year

4. *Naphthol AS-D chloroacetate

5. *Buffered staining solution-------- 1 hour

6. Lillie-Mayer hematoxylin -------- 1 year

Oil Red O (fats)

1. Oil red O, stock -------- 1 year

2. Oil red O, working -------- 1 year

3. Lillie-Mayer hematoxylin -------- 1 year

4. Glycerin-jelly -------- 10 years

Sudan Black B (phospholipids)

1. Sudan black B -------- 1 year

2. Nuclear fast red -------- 4 months

3. Glycerin-jelly -------- 10 years

Hematoxylin and Eosin (H&E)

1. Harris hematoxylin -------- 6 months

2. Lillie-Mayer hematoxylin -------- 1 year

3. Alcoholic eosin, stock -------- 1 year

4. Alcoholic eosin, working -------- 6 months

Strong-Fast (smears and touch preparations)

1. Fixative -------- 1 year

2. Phosphate buffer, pH 6.8 -------- 1 year

3. 10% Triton X-100 -------- 6 months

4. Solution I (eosin Y) -------- 1 year

5. Solution II (azure A-methylene blue) -------- 1 year