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Bensley’s trichrome for Muscle and Collagen

Bensley's trichrome

for Muscle and Collagen



Solution A

Acid fuchsinTo saturation
Aniline water100mL

Solution Preparation

  1. Add about 20 g dye to the aniline water.
  2. Shake intermittently for 48 hours. Filter.

Solution B

Phosphomolybdic acid1g
Distilled water100mL

Solution C

Orange G2g
Aniline blue0.5g
Distilled water100mL

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.


  1. Bring sections to water via xylene and ethanol.
  2. Place into solution A for 10 minute.
  3. Rinse with distilled water.
  4. Place into solution B for 10 minutes.
  5. Rinse quickly with distilled water.
  6. Place into solution C for 60 minutes.
  7. Rinse with 95% ethanol until clouds of dye cease being extracted.
  8. Dehydrate with ethanol.
  9. Clear with xylene and mount with a resinous medium.

Expected Results

  • Nuclei  –  red
  • Erythrocytes  –  orange
  • Cytoplasm  –  red
  • Collagen  –  blue


  • Solution A is Altmann’s acid fuchsin, originally formulated for staining mitochondria.
  • This method does not specify an acid resistant nuclear stain. Doing so before staining with solution A may improve the technique.

Safety Note

Prior to handling any chemical, consult the Safety Data Sheet (SDS) for proper handling and safety precautions.


  1. Gray, Peter. (1954)
    The Microtomist’s Formulary and Guide.
    Originally published by: The Blakiston Co.
    Republished by: Robert E. Krieger Publishing Co.
    Mallory, F. B., (1938),
    Pathological technique.
    Saunders, Philadelphia, Pa, USA