Harris'
Alum Hematoxylin

Ingredient Amount Function
Hematoxylin 5 g Dye
Potassium alum 100 g Mordant
Distilled water 1 L Solvent
95% ethanol 50 mL Solvent
Mercuric oxide 2.5 g Oxidant
Glacial acetic acid 40 mL Acidifier

 

Compounding procedure
Dissolve the hematoxylin in the ethanol.
Dissolve the alum in warmed distilled water.
Combine the solutions in an oversize container.
Bring to the boil.
Add the mercuric oxide, remove from heat.
Cool rapidly.
Add the glacial acetic acid.
It may be used immediately when cool.

Method

  1. Bring sections to water with xylene and ethanol.
  2. Paraffin sections should be stained for 5 minutes, and cytology smears for 45 seconds.
  3. Rinse well with water.
  4. Differentiate with acid ethanol if necessary.
  5. Rinse with water and blue.
  6. Rinse well with water.
  7. Counterstain if desired.
  8. Dehydrate with ethanol, clear with xylene and mount with a resinous medium.

Expected results

Notes

  1. The solution is a regressive formulation, although it is sometimes used progressively with short staining times.
  2. When the mercuric oxide is added, the solution may foam.
  3. Mercuric oxide is toxic and no longer recommended for oxidation of hematoxylin. If used, all discarded solution should be disposed of as mercury contaminated. Sodium iodate is sometimes substituted, based on a maximum of 0.2 gram sodium iodate for each 1 gram of hematoxylin. In practice, 0.5 gram works quite well.
  4. The acetic acid is optional, but nuclear staining is more precise if it is included.
  5. A precipitate often forms, so the solution should be filtered before use.
  6. As with many strong hematoxylin solutions, mucin may be blue.
  7. Acid ethanol is 0.5% - 1% hydrochloric acid in 70% ethanol.
  8. Blueing is done with alkaline solutions such as hard tap water, Scott’s tap water substitute, 0.1% ammonia water, 1% aqueous sodium acetate, 0.5% aqueous lithium carbonate etc.

 

Reference
Culling, C.F.A., Allison, R.T. and Barr, W.T.
Cellular Pathology Technique, Ed.4.
Butterworth, London, UK.

Drury, R.A.B. and Wallington, E.A., (1980)
Carleton's histological technique Ed. 5
Oxford University Press, Oxford, UK.

Bancroft, J.D. and Stevens A. (1982)
Theory and practice of histological techniques Ed. 2
Churchill Livingstone, Edinburgh & London, UK.

 


 

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