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Dahlia-Iron-Resorcin for Elastic Fibres

Dahlia-Iron-Resorcin

for Elastic Fibres

8
steps
9
materials

Materials

    • Weigert’s iron hematoxylin or equivalent
    • Van Gieson’s picro-fuchsin
    • Iron-resorcin-dahlia solution
      MaterialAmount
      Dahlia2g
      Resorcin4g
      Ferric nitrate, 30% aqu.50mL
      0.5% aqu. hydrochloric acid200mL
      Methanol180mL
      Acetone20mL
      Hydrochloric acid, conc.4mL

Preparation

  1. Bring 200 mL of 0.5% hydrochloric acid in an oversize flask to the boiling point, remove from the heat and carefully add the dahlia.
  2. Return to the boil, and add the resorcin and ferric nitrate.
  3. Continue boiling for 3 minutes, then cool and filter. Discard the filtrate. Dry the filter paper and beaker, then place the precipitate and filter paper back into the flask.
  4. Add the methanol, acetone and hydrochloric acid.
  5. Heat carefully on a hot plate until the precipitate dissolves.

Tissue Sample

5µ paraffin sections of neutral buffered formalin fixed tissue are suitable. Other fixatives are likely to be satisfactory.

Protocol

  1. Bring sections to water via xylene and ethanol.
  2. Do a Mallory bleach.
  3. Place into the staining solution for 30 minutes or longer.
  4. Wash with 95% ethanol to remove excess solution.
  5. Differentiate with 1% acid alcohol if necessary.
  6. Wash in water.
  7. Counterstain with iron hematoxylin and van Gieson.
  8. Dehydrate with ethanol, clear with xylene and mount with a resinous medium.

Expected Results

  • Elastic fibres  –  blue black
  • Collagen  –  red
  • Cytoplasm  –  yellow
  • Nuclei  –  blue

Notes

  • Between the 1st and 2nd editions of their book, Bancroft and Stevens changed the solvent for the precipitate to the following:
    SolventAmount
    2-methoxyethanol100mL
    Distilled water100mL
    Hydrochloric acid, conc.4mL

    Dry the filter paper and beaker, then place the precipitate and filter paper back into the flask. Add the mixture and heat carefully on a hot plate until the precipitate dissolves.

  • 2-methoxyethanol is also known as ethylene glycol monomethyl ether.
  • The authors suggest the following dyes as suitable substitutes for dahlia:
    • Methyl violet
    • Ethyl violet
    • Victoria blue
    • Thionin
  • Note that this method uses ferric nitrate instead of ferric chloride.

Safety Note

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

References

  1. Bancroft, J. D. and Stevens, A.,
    Theory and Practice of Histological Techniques, Ed. 1 (1977),
    Churchill Livingstone, London, UK.
  2. Bancroft, J. D. and Stevens, A.,
    Theory and Practice of Histological Techniques, Ed. 2 (1982),
    Churchill Livingstone, London, UK.