Koneff's Trichrome
for Pituitary Cells

Solutions

Solution A
Aniline 0.1 mL
Ethanol, 90% 100 mL
Solution B
Acetic acid, glacial 1 mL
Ethanol, 90% 100 mL
Solution C
Azocarmine 1 g
Acetic acid, glacial 1 g
Distilled water 100 mL
Solution D
Aniline 0.06 mL
Ethanol, 90% 100 mL
Solution E
Phosphotungstic acid 5 g
Distilled water 100 mL
Solution F
Aniline blue 0.5 g
Orange G 2 g
Oxalic acid 2 g
Phosphotungstic acid 0.05 g
Distilled water 100 mL
Solution G
Acetic acid 1 mL
Distilled water 100 mL

Tissue sample
The instructions specify fixation in Baley's fixative (see notes). Other mercuric-chrome fixatives would probably be satisfactory. The instructions also specify 3-4 µ celloidin sections which have been attached to a slide, and from which the celloidin has been removed. Possibly, similar paraffin sections would be satisfactory.

Method

  1. Attach sections to slides and remove celloidin, or
    Dewax sections with xylene and bring to ethanol.
  2. Although not specified, a water rinse followed by the iodine thiosulphate sequence and a second water wash, to remove mercury pigment would be advantageous at this point.
  3. Wash with 70% ethanol.
  4. Place into solution A for 45 minutes.
  5. Place into solution B for 1-2 minutes.
  6. Place into solution C for 2 hours at 56°C.
  7. Wash with water.
  8. Place into solution D until nuclei are red and the cytoplasm is pink.
  9. Place into solution B for 1-2 minutes.
  10. Place into solution E for 4 hours
  11. Place into solution F for 4 hours until basophils are blue.
  12. Return to solution E for 3-5 minutes.
  13. Wash with water.
  14. Rinse with solution G.
  15. Wash with water.
  16. Dehydrate with absolute ethanol.
  17. Clear with xylene and mount with a resinous medium.

Expected results

Notes

  1. Baley's fixative consists of:
    Mercuric chloride 3.4 g
    Potassium dichromate 3.4 g
    Concentrated formalin (40%) 25 mL
    Water 225 mL
    Return to fixation instructions.

 

Reference

Gray, Peter. (1954)
The Microtomist's Formulary and Guide.
Originally published by:– The Blakiston Co.
Republished by:– Robert E. Krieger Publishing Co.
  Citing:–
    Koneff, (1938)
    Stain Technology, v.13, pp.49

 


 
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Last updated May 2005.