McFarlane's Trichrome 1st Variant
for Muscle and Collagen
Materials
- Weigert’s iron hematoxylin or equivalent
- Solution A
Material Amount Picric acid 0.2 g Acid fuchsin 1 g Aniline blue 2 g Phosphotungstic acid 1 g Acetic acid, glacial 2 mL Distilled water 98 mL - Solution B
Material Amount Acetic acid, glacial 2 mL Distilled water 100 mL - Solution C
Material Amount Picric acid 0.25 g Phosphotungstic acid 2.5 g Ethanol, 95% 10 mL Distilled water 90 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. Formalin fixed tissues may benefit from secondary fixation of sections in Bouin’s fluid.
Protocol
- Bring sections to water via xylene and ethanol.
- Stain nuclei with Weigert’s iron hematoxylin or equivalent.
- Place into solution A for 5 minute.
- Rinse with solution B.
- Rinse with distilled water.
- Place into solution C until differentiated.
- Rinse with distilled water.
- Wash with solution B.
- Dehydrate with ethanol.
- Clear with xylene and mount with a resinous medium.
Expected Results
- Nuclei – black
- Erythrocytes – yellow
- Muscle – red
- Cytoplasm – red
- Collagen – blue
Notes
- McFarlane also published two other, multi-step, trichrome methods. One is simpler than the other.
- Solutions A and C require dry weights of picric acid. This could be provided from a saturated aqueous solution (1 g in 78 mL). Alternate formulas are:
Solution A
Material Amount Picric acid, sat. aqueous 16 mL Distilled water 82 mL Acetic acid, glacial 2 mL Acid fuchsin 1 g Aniline blue 2 g Phosphotungstic acid 1 g Solution C
Material Amount Picric acid, sat. aqueous 20 mL Distilled water 70 mL Ethanol, 95% 10 mL Phosphotungstic acid 2.5 g
Safety Note
Prior to handling any chemical, consult the Safety Data Sheet (SDS) for proper handling and safety precautions.
References
- Gray, Peter. (1954)
The Microtomist’s Formulary and Guide.
Originally published by: The Blakiston Co.
Republished by: Robert E. Krieger Publishing Co.
Citing:
McFarlane, (1944)
Stain Technology, v. 19, pp. 29
