Debiden's Alum Hematoxylin Variants
Materials
Material | Variant | Function | |
---|---|---|---|
1987 | 1991 | ||
Hematoxylin | 5 g | 5 g | Dye |
Potassium alum | 100 g | 100 g | Mordant |
Distilled water | 1 L | 1 L | Solvent |
Glacial acetic acid | 20 mL | 20 mL | Acidifier |
Mercuric oxide, red | 2.5 g | – | Oxidant |
Javex (5.25% sodium hypochlorite) | – | 2 mL | Oxidant |
Compounding procedures
- Dissolve the alum in the water in a large flask.
- Add the hematoxylin. Mix well.
- Place in a 60-65°C oven overnight.
- Remove from the oven, and add the appropriate oxidizing agent.
- Stir until cold. Filter.
Preparation notes
1980
- Acetic acid may be added as soon as it is cooled following preparation.
- It is usable immediately.
- It is preferable to prepare small volumes monthly.
1991
- This improves in staining ability if allowed to ripen for a week or two.
- If acetic acid is used, it should be added just before use.
Protocol
- Bring sections to water with xylene and ethanol.
- Place into the staining solution for an appropriate time.
- Rinse well with water.
- Differentiate with acid ethanol if necessary.
- 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
- These two solutions are modifications of Harris’ hematoxylin and are regressive.
- Paraffin sections should be stained for 5 minutes, and cytology smears for 45 seconds.
- As with many strong regressive formulae, mucin may be stained blue.
- Acid ethanol is 0.5% – 1% hydrochloric acid in 70% ethanol.
- 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
- Debiden, D., (1987)
Improved preparation of Harris’ hematoxylin
Histologic, v.18, No.8 - Debiden, D., (1991)
A new oxidant for Harris’ hematoxylin
Histologic, v.21, No.2