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Birch-Hirschfeld’s Crystal Violet for Amyloid

Protocol

Birch-Hirschfeld's Crystal Violet

for Amyloid

9
steps
4
materials
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Materials

Tissue Sample

Frozen sections are preferred. Cryostat sections usually show brighter metachromasia. Unmounted frozen sections may also be floated in each solution and mounted on a slide just before coverslipping. 5µ paraffin sections of neutral buffered formalin fixed tissue are likely also suitable. Other fixatives may be satisfactory.

Protocol

  1. Bring sections to water via xylene and ethanol, except for cryostat and frozen sections.
  2. Place into bismarck brown solution for 5 minutes.
  3. Rinse well with 95% ethanol, then rinse with distilled water.
  4. Place into crystal violet solution for 5 minutes.
  5. Rinse with water.
  6. If necessary, differentiate in 1% acetic acid until amyloid is red and contrasts well with the tissue.
  7. Wash well in tap water.
  8. Drain all water from the slide until just damp and mount with levulose syrup.
  9. Ring the coverslip to inhibit evaporation of the mounting medium.

Expected Results

  • Amyloid  –  purple-red
  • Background  –  blue-violet
  • Nuclei  –  brown

Notes

  • Methyl violet may be used instead of crystal violet if preferred.
  • Although levulose syrup (fructose syrup or high fructose corn syrup) is specified it is likely that Highman’s gum syrup would be preferable as it inhibits leaching of the dye.
  • Although bismarck brown is metachromatic (yellow metachromasia), it is used here as a basic dye for staining nuclei.

Safety Note

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

References

  1. Gray, Peter. (1954)
    The Microtomist’s Formulary and Guide, p.451.
    Originally published by: The Blakiston Co.
    Republished by: Robert E. Krieger Publishing Co.