Hematoxylin Stain

Solutions

Hematoxylin

Hematoxylin	6.4g
Ammonium alum	60g
Ethanol	200ml
Glycerol	160ml
Distilled water	640ml

Mix for a day then allow the solution to ripen in the dark for at least a month before using.

Acidic Alcohol

Concentrated HCl	4ml
95% Ethanol	396ml

Bluing Agent

Sodium bicarbonate	1g
Distilled water	1 l

Staining

1. Deparaffin and rehydrate slides.
2. Slightly overstain the sections with hematoxylin 3-5 minutes.
3. Remove excess stain in tap water for 2 minutes.
4. Differentiate and restain a few seconds in acidic alcohol until sections look red.
5. Rinse in tap water.
6. Blue in bicarbonate until nuclei stand out sharply blue for 2 minutes.
7. Rinse in running tap water.
8. Dehydrate and clear, coverslip with Cytoseal.