Musto's Hematoxylin
for Elastic Fibres


Stock hematoxylin
Hematoxylin 2 g
Ethanol, 95% 100 mL
Stock ferric chloride
Ferric chloride, hexahydrate 12.4 g
Distilled water 500 mL
Hydrochloric acid, concentrated 5 mL
Stock iodine
Potassium iodide 4 g
Iodine 2 g
Distilled water 100 mL
Thiosulphate solution
Sodium thiosulphate 5 g
Distilled water 100 mL
Working solution
Stock hematoxylin 30 mL
Stock ferric chloride 20 mL
Stock iodine 10 mL

Tissue sample
A 10% Formalin variant is suitable.
Paraffin sections at 5µ are preferred.


  1. Bring sections to water via xylene and ethanol.
  2. Place sections in Bouin's fluid at 60°C for 1 hour.
  3. Remove the yellow colouration with running tap water.
  4. Place in working solution for 45 minutes.
  5. Wash well with tap water, then rinse with distilled water.
  6. Place into thiosulphate solution for 2 minutes to remove iodine discolouration.
  7. Wash well with tap water, then rinse with distilled water.
  8. Counterstain with Van Gieson.
  9. Dehydrate with absolute ethanol.
  10. Clear with xylene and mount with a resinous medium

Expected results


  1. The original paper recommended counterstaining with a modification of the HPS.
  2. The treatment with Bouin's solution is likely included to improve an HPS stain. It may be possible to eliminate it for counterstaining with Van Gieson.
  3. This is a modification of Verhöeff's hematoxylin which does not require differentiating, although the Van Gieson counterstain may diminish the intensity of the stain.


Musto, Linda, (1981),
Improved iron-hematoxylin stain for elastic fibers.,
Stain Technology, v 56, page 185-7.




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