Kefalas's Iron Hematoxylin

Solution A Amount Function
Hematoxylin 1 g Dye
Ferric chloride 1 g Mordant
Acetone 100 mL Solvent
Hydrochloric acid 0.05 mL Acidifier


Compounding procedure
Dissolve the ferric chloride and hematoxylin in the acetone.
Add the hydrochloric acid.


  1. Bring sections to water with xylene and ethanol.
  2. Place into the staining solution for 30 minutes.
  3. Wash well in running tap water to blue.
  4. Rinse with distilled water.
  5. Counterstain if desired.
  6. Dehydrate with ethanol, clear with xylene and mount with a resinous medium.

Since this solution is almost anhydrous it can be used in a staining procedure which completely avoids water or ethanol.

  1. Remove wax with xylene.
  2. Remove xylene with a few changes of acetone.
  3. Place into staining solution for 30 minutes.
  4. Wash with acetone.
  5. Clear with xylene and mount with a resinous medium.

Expected results


  1. 0.05 mL hydrochloric acid would be about 1-2 drops.
  2. The simplicity of the formula indicates it is likely not stable for long.
  3. The presence of hydrochloric acid may indicate progressive staining.
  4. It is likely suitable as an acid resistant nuclear stain.
  5. Although the staining time should be established by trial and error. 30 minutes would likely suffice as a starting point.


Gray, Peter. (1954)
The Microtomist's Formulary and Guide.
Originally published by:– The Blakiston Co.
Republished by:– Robert E. Krieger Publishing Co.
    Kefalas, (1926), Journal of the Royal Microscopical Society v. 46, p. 277.



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