SURGICAL PATHOLOGY - HISTOLOGY Date: STAINING MANUAL - CONNECTIVE TISSUE Page: 1 of 4 PURPOSE: Ud to differentiate between collagen and smooth muscle in tumors, and the increa of collagen in dias such as cirrhosis. Routine stain for liver and kidney biopsies.
PRINCIPLE: As the name implies, three dyes are employed lectively staining muscle, collagen fibers, fibrin, and erythrocytes. The general rule in trichrome staining is that the less porous tissues are colored by the smallest dye molecule; whenever a dye of large molecular size is able to penetrate, it will always do so at the expen of the smaller molecule. Others suggest that the tissue is stained first with the acid dye, Biebrich Scarlet, which binds with the acidophilic tissue components. Then when treated with the phospho acids, the less permeable components retain the red, while the red is pulled out of the collagen. At the same time causing a link with the collagen to bind with the aniline blue.
FIXATIVE: Bouin's is preferred, 10% formalin.
TECHNIQUE: Cut paraffin ctions 4m
EQUIPMENT: Rin glassware in DI water. Coplin jars, 60°C oven or waterbath, microwave
REAGENTS:
Bouin’s Fixative乌兰木图山
Saturated picric acid1500.0 ml Formaldehyde500.0 ml Glacial acetic acid100.0 ml Mix well, label, date and initial. Stable for 2 years.
CAUTION: Carcinogenic, irritant.Biebrich Scarlet:
Biebrich scarlet 2.7 gm Acid fuchsin0.3 gm Distilled water300.0 ml Glacial acetic acid 3.0 ml Mix well, label with initial and date. Solution is stable for 6 months.
CAUTION: Avoid contact and inhalation.
MASSON’S TRICHROME Page: 2 of 4
Weigert's Iron Hematoxylin Stock Solution A:三年级数学竖式
Hematoxylin 5.0 gm 95% alcohol500.0 ml Mix well, label with initial and date. Stable for 1 year.
CAUTION: Flammable,avoid contact and inhalation.
Stock Solution B:
29% ferric chloride20.0 ml Distilled water475.0 ml Hydrochloric acid 5.0 ml Mix well, label with initial and date. Stable for 1 year.
CAUTION: Corrosive, avoid contact and inhalation.
Weigert's Hematoxylin
Working Solution
Solution A25.0 ml Solution B25.0 ml Mix well, solution will remain stable for 3 - 4 days.Phosphotungstic/ Phosphomolybdic Acid Solution: Phosphotungstic acid 25.0 gm Phosphomolybdic acid25.0 gm Distilled water1000.0 ml Mix well, label with initial and date. Solution is stable for 6 months.
CAUTION: Avoid contact and inhalation. Aniline Blue:
Aniline blue 2.5 gm Distilled water100.0 ml Glacial acetic acid 1.0 ml Mix well, label with initial and date. Solution is stable for 6 months.
Caution: Avoid contact and inhalation. 1% Acetic Acid:
Glacial acetic acid10.0 ml Distilled water1000.0 ml Mix well, label with initial and date. Solution is stable for 1 year. Caution: Avoid contact and inhalation.
SAFETY: Wear gloves, goggles and lab coat. Avoid contact and inhalation of dyes and chemicals.
Bouin's contains: formaldehyde, a known carcinogen, picric acid can become explosive when dry. Toxic by skin absorbtion. Keep hot uncapped Bouin’s under the hood.
Phosphomolybdic, phosphotungsic acid powders, and acetic acid solutions are skin and eye irritants, and strong corrosives.
MASSON’S TRICHROME Page: 3 of 4 1.*Mordant in Bouin's solution, microwave 1 minute, allow to stand 15
minutes.
2.Wash in running tap water to remove the picric acid, 5 minutes.
3.Weigert's working hematoxylin, 10 minutes.
4.Blue in running tap water for 5 minutes, rin in distilled water.
湖南怀化学院
5.Biebrich scarlet for 5 minutes.
6.Rin in distilled water.
7.Phosphotungstic/phosphomolybdic acid for 10 minutes, discard
solution.
8.Transfer directly into Aniline blue for 5 minutes.
9.Rin in distilled water.
10.1% Acetic acid for 1 minute, discard solution, rin in distilled water.
11.Dehydrate, clear, and coverslip.
*Conventional method: Mordant in Bouin's solution, 60°C for 1 hour. RESULTS:
Nuclei black开学搞笑图片
Cytoplasm, muscle, erythrocytes red
Collagen blue
NOTES:
1.Light green may be substituted for Aniline blue..
2.5% phosphotungstic acid for 5 minutes, must be substituted when
using Light green.
3.When staining liver biopsies, the collagen is better light blue than
dark blue.
REFERENCE:
Sheehan D, Hrapchak B, Theory and practice of Histotechnology, 2nd Ed, 1980, pp 189-190, Battelle Press, Ohio
Bancroft J, Stevens A, Theory and Practice of Histological Techniques, 2nd Ed, 1982, pp 131-135, Churchill-Livingston, NY
Luna L, Manual of Histologic Staining Methods of the AFIP, 3rd Ed, 1968, pp 94-95, McGraw-Hill, NY
Carson F, Histotechnology A Self-Instructional Text, 1st Ed, 1990, pp 142-143, ASCP, Ill
咋怎么写CONNECTIVE TISSUE
MASSON’S TRICHROME Page: 4 of 4 Laboratory, 2nd ED, 1991, Anatech
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TRICHROME STAIN, MASSON'S (TRI)-COLLAGEN Page 1 of 2
1.*Mordant in Bouin's solution, microwave 1 minute, allow to stand 15 minutes.
2.Wash in running tap water to remove the picric acid, 5 minutes.
蔽的组词
3.Weigert's working hematoxylin, 10 minutes.
4.Blue in running tap water for 5 minutes, rin in distilled water.
5.Biebrich scarlet for 5 minutes.
6.Rin in distilled water.
7.Phosphotungstic/phosphomolybdic acid for 10 minutes, discard solution.
8.Transfer directly into Aniline blue for 5 minutes.
9.Rin in distilled water.
10.1% Acetic acid for 1 minute, discard solution, rin in distilled water.
11.Dehydrate, clear, and coverslip.
*Conventional method: Mordant in Bouin's solution, 60°C for 1 hour.
RESULTS:
Nuclei black
Cytoplasm, muscle, erythrocytes red
Collagen blue
SAFETY/PPE: Wear gloves, goggles and lab coat. Keep hot Bouin’s under the fume hood.
Weigert's Iron Hematoxylin
Stock Solution A:
Hematoxylin 5.0 gm 95% alcohol500.0 ml Mix well, label with initial and date. Stable for 1 year.
CAUTION: Flammable
Stock Solution B:
29% ferric chloride20.0 ml Distilled water475.0 ml Hydrochloric acid 5.0 ml Mix well, label with initial and date. Stable for 1 year.
Working Solution
Solution A25.0 ml Solution B25.0 ml Mix well, solution will remain stable for 3 -4 days.Bouin’s Fixative
CAUTION: Carcinogenic, irritant, toxic.
Biebrich Scarlet:
Biebrich scarlet 2.7 gm Acid fuchsin0.3 gm Distilled water300.0 ml Glacial acetic acid 3.0 ml Mix well, label with initial and date. Solution is stable for 6 months.
CAUTION: Possible carcinogen Phosphotungstic/
Phosphomolybdic Acid Solution: Phosphotungstic acid 25.0 gm Phosphomolybdic acid25.0 gm Distilled water1000.0 ml Mix well, label with initial and date. Solution is stable for 6 months.
TRICHROME STAIN, MASSON'S (TRI)-COLLAGEN Page 2 of 2
Aniline B lue:
苹果5s怎么刷机Aniline blue 2.5 gm Distilled water100.0 ml Glacial acetic acid 1.0 ml Mix well, label with initial and date. Solution is stable for 6 months.1% Acetic Acid:
Glacial acetic acid10.0 ml Distilled water1000.0 ml Mix well, label with initial and date. Solution is stable for 1 year.
