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Smith’s Vanadium Hematoxylin

Smith's Vanadium Hematoxylin

7
steps
5
materials

When differentiated with dilute lithium carbonate, this solution demonstrates basic proteins, including nuclear histones.

Materials

MaterialAmountFunction
Hematoxylin50 mgDye
Ethanol, 100%5 mLSolvent
Glycerol10 mLSolvent
Distilled water35 mLSolvent
Ammonium metavanadate200 mgSolvent

Compounding procedure

  1. Dissolve the hematoxylin in the ethanol, then add the glycerol and water in sequence.
  2. Add the ammonium vanadate and stir for 30 minutes.
  3. The ammonium vanadate may not completely dissolve. The pH is 6.2.

Tissue Sample

Paraffin sections of Schaudinn, Bouin or formalin fixed tissues are suitable. Sections of Zenker or Helly fixed tissues must be soaked in saturated aqueous lithium carbonate for 4 hours to stain satisfactorily.

Protocol

  1. Bring sections to water with xylene and ethanol.
  2. Place into the vanadate hematoxylin for 30 minutes.
  3. Place directly into 0.08% aqueous lithium carbonate for the appropriate time.
    Differentiation times in lithium carbonate

    FixativeTime
    Bouin2 minutes
    Formalin, 10%2 minutes
    Helly10 minutes
    Schaudinn5 minutes
    Zenker10 minutes
  4. Briefly rinse with distilled water for 2 seconds.
  5. Optionally, counterstain if wished.
  6. Dehydrate with 70%, 95% and absolute ethanols.
  7. Clear with xylene and mount with a resinous medium (Permount specified).

Expected Results

  • Nuclear histone  –  blue
  • Ribosomal protein  –  paler blue
  • Other protein  –  as counterstain or unstained

Notes

  • Counterstaining can be accomplished at step 5 with 1% aqueous eosin Y, phloxine B or erythrosin B.
  • Alternatively, metanil yellow at 0.1% in 95% ethanol containing 0.1% acetic acid may be used between the 70% and 95% ethanols in step 6.
  • Counterstaining with 0.01% aqueous safranin O stained RNA intensely, giving good contrast between blue nuclear histone and endoplasmic reticulum.

Safety Note

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

References

  1. Smith, A. A., (1995)
    A vanadate hematoxylin stain for basic proteins.
    Biotechnic and Histochemistry, v. 70, Nº 5, p. 5.