Horneyold's Alum Hematoxylin
Materials
Hematoxylin Solution
Material | Amount | Function |
---|---|---|
Hematoxylin | 0.7 g | Dye |
100% ethanol | 20 mL | Solvent |
Alum Solution
Material | Amount | Function |
---|---|---|
Ammonium alum | 0.35 g | Mordant |
Distilled water | 60 mL | Solvent |
Tincture of Iodine
Material | Amount | Function |
---|---|---|
Iodine | 70 g | Oxidant |
Potassium iodide | 50 g | – |
Distilled water | 50 mL | Solvent |
100% ethanol | 950 mL | Solvent |
Compounding Procedures
Tincture of iodine
- Mix the dry iodine and potassium iodide together.
- Add the water and mix well to dissolve.
- Add the ethanol and mix well.
Staining solution
- Dissolve the hematoxylin in the ethanol and the Alum in the water.
- Combine the two solutions and mix well.
- Leave in the light by a window for 3-4 days.
- Add 20 drops tincture of iodine.
Protocol
- Bring sections to water with xylene and ethanol.
- Place into the staining solution for 5-10 minutes.
- Rinse well with water.
- Differentiate with acetic ethanol.
- Rinse with water and blue.
- Rinse well with water.
- Counterstain if desired.
- Dehydrate with ethanol, clear with xylene and mount with a resinous medium.
Expected Results
- Nuclei – blue
- Background – as counterstain or unstained
Notes
- Horneyold’s formula is a modification of Böhmer’s Alum hematoxylin.
- This solution is regressive, and is recommended for osmium fixed tissue.
- Acetic ethanol is 70% ethanol containing glacial acetic acid. The amount of acetic acid to add was not given, but 0.5%-1% should suffice.
- 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.
Safety Note
Prior to handling any chemical, consult the Safety Data Sheet (SDS) for proper handling and safety precautions.
References
- Bolles Lee, A.. Edited by Gatenby, J.B. and Beams, H.W., (1950)
The Microtomist’s Vade-Mecum. 11 ed.,
Churchill, London, UK. - Lange, N.A., (1966)
Lange’s Handbook of Chemistry, Revised 10th ed.,
McGraw-Hill.