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Heidenhain’s Iron Hematoxylin

Heidenhain's Iron Hematoxylin

8
steps
4
materials

Materials

Solution A

MaterialAmountFunction
Ferric ammonium sulfate2.5 gMordant
Distilled water100 mLSolvent

Solution

MaterialAmountFunction
Hematoxylin0.5 gDye
Distilled water100 mLSolvent

Compounding Procedure

  1. Make each solution separately.
  2. Solution B should be ripened for a minimum of one month.

Protocol

  1. Bring sections to water with xylene and ethanol.
  2. Place into solution A for 30 minutes to 24 hours.
  3. Rinse with distilled water.
  4. Place into solution B for 30 minutes to 24 hours.
  5. Rinse with tap water.
  6. Differentiate in solution A, controlling microscopically.
  7. Wash well in running tap water to blue.
  8. Dehydrate with ethanol, clear with xylene and mount with a resinous medium.

Expected Results

  • Nuclei and other structures  –  blue to black

Notes

  • The stock solutions are stable for some time.
  • The hematoxylin solution needs to be ripened.
  • Staining at elevated temperatures (not over 60°C) will shorten the required times. Always differentiate at room temperature.
  • The degree of differentiation will determine which tissue components are prominent. The method can demonstrate many structures, including chromosomes, nuclear components, mitochondria and muscle striations
  • The solutions may be reused, with the exception of solution A, which is used to differentiate, which should be fresh each time.
  • Counterstaining is not recommended.
  • This method is usually recommended for monochrome photography.
  • Compare Heidenhain’s iron hematoxylin technique to other published variations.

Safety Note

Prior to handling any chemical, consult the Safety Data Sheet (SDS) for proper handling and safety precautions.

References

  1. Gray, Peter. (1954)
    The Microtomist’s Formulary and Guide.
    Originally published by: The Blakiston Co.
    Republished by: Robert E. Krieger Publishing Co.
    Citing:
    Heidenhain, M., (1892).
    Festschrift Herrn A. von Kolloker zur Feier seines
    fünfzigjährigen medicinischen
    Doktorjubiläums, p.118.
    Wilhelm Engellmans, Leipzig, Germany