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Bencosme’s Alum Hematein

Protocol

Bencosme's Alum Hematein

8
steps
8
materials
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Materials

Original

MaterialAmountFunction
Hematein2 gDye
Potassium alum133 gMordant
Distilled water920 mLSolvent
Glacial acetic acid20 mLAcidifier

Variant

MaterialAmountFunction
Hematein2.5 gDye
Potassium alum120 gMordant
Distilled water1 LSolvent
Glacial acetic acid10 mLAcidifier

Compounding procedure

  1. Add the alum to the water in a 2 L flask and bring to a boil for 1 minute.
  2. Add the hematein and mix by swirling.
  3. Place an inverted funnel over the flask and simmer for 10 minutes, shaking frequently.
  4. Cool to room temperature and add the acetic acid.
  5. Filter before use and twice weekly.
  6. It may be used immediately.

Protocol

  1. Bring sections to water with xylene and ethanol.
  2. Place into the staining solution for 2 minutes.
  3. Rinse with water.
  4. Differentiate with 0.5% hydrochloric acid in 70% ethanol for one second.
  5. Rinse well with water and blue.
  6. Rinse well with water.
  7. Counterstain with HPS.
  8. Dehydrate with ethanol, clear with xylene and mount with a resinous medium.

Expected Results

  • Nuclei  –  blue
  • Background  –  shades of red and yellow.

Notes

  • Note that this solution uses hematein rather than hematoxylin. Hematoxylin could be used, but would require 0.4 grams sodium iodate to be added along with the dye for complete oxidation in the original solution, although 0.3 grams might be more appropriate to extend the life. Comparable amounts would be 0.5 g and 0.4 g, respectively, for the variant formula.
  • When freshly made this solution stains in 2 minutes. Staining time slowly increases to 10 minutes as the solution ages. Life is about a month.
  • 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

  1. Medical Laboratory Technology, 2nd ed. (1969)
    Lynch M.J., Raphael S.S., Mellor L.D., Spare P.D. and Inwood M.J.H.,
    W. B. Saunders Co., Toronto, On., Canada