Crossman's Trichrome
for Muscle and Collagen

Solutions

Solution A
Acid fuchsin	0.3	g
Orange G	2	g
Acetic acid, glacial	1	mL
Distilled water	100	mL
Thymol	a	small crystal

Solution B
Phosphomolybdic acid	1	g
Distilled water	100	mL

Solution C
Either				or
Aniline blue	2	g		Light green SF	1	g
Acetic acid, glacial	2	mL		Acetic acid, glacial	1	mL
Distilled water	98	mL		Distilled water	99	mL

Solution D
Acetic acid, glacial	1	mL
Distilled water	99	mL

Tissue sample
5µ paraffin sections of neutral buffered formalin fixed tissue are suitable. Other fixatives are likely to be satisfactory. Most trichrome stains benefit from picric acid or mercuric chloride fixation.

Method

1. Bring sections to water via xylene and ethanol.
2. Stain nuclei with an acid resistant nuclear stain such as Weigert's iron hematoxylin.
3. Place into solution A for 1 minute.
4. Rinse with distilled water.
5. Place into solution B until collagen is decolorised.
6. Rinse quickly with distilled water.
7. Place into solution C for 5 minutes.
8. Rinse with distilled water.
9. Place into solution D until differentiated.
10. Rinse with distilled water.
11. Dehydrate with ethanol.
12. Clear with xylene and mount with a resinous medium.

Expected results

• Nuclei  –  black
• Erythrocytes  –  yellow
• Cytoplasm  –  red
• Collagen  –  blue or green

Reference

Gray, Peter. (1954)
The Microtomist's Formulary and Guide.
Originally published by:– The Blakiston Co.
Republished by:– Robert E. Krieger Publishing Co.

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