Lendrum, Slidders and Fraser's
Trichrome for Collagen and Other Tissues

Dye CI name Author's Code
 
Small molecular weight dyes
Martius yellow Acid yellow 24 AY24
Orange G Acid orange 10 A010
Azo-eosin Acid red 4 AR4
Propalan red 3GX Acid red 57 AR57
Acid fuchsin Acid violet 19 AV19
 
Large molecular weight dyes
Sun yellow Direct yellow 11 RY11
Sirius red F3B Direct red 80 DR80
Benzo new blue GS Direct blue 10 DR10
Durazol brilliant blue B Direct blue 109 DB109

Solutions

Small MW solution
Dye 0.5 g
Phosphotungstic acid 2 g
Ethanol, 95% 100 mL
Large MW solution
Dye 0.5 g
Acetic acid, glacial 1 mL
Distilled water 99 mL
Use one of the dyes from the appropriate list above for each solution.

Tissue sample
3-5µ paraffin sections of formal sublimate fixed tissue is preferred. If formalin fixation is used, the sections may require secondary fixation with picro-mercuric-alcohol.

Method

  1. Bring sections to water via xylene and ethanol.
  2. Stain nuclei with an acid resistant nuclear stain.
  3. Rinse with 95% ethanol.
  4. Place into small MW solution for 3 minutes.
  5. Rinse with distilled water.
  6. Place in the large MW solution for 30 minutes to 2 hours.
  7. Rinse with water.
  8. Dehydrate with absolute ethanol.
  9. Clear with xylene and mount with a resinous medium.

Expected results
The results vary depending on the particular combination of small and large molecular weight dyes chosen. As a general guide, the small molecular weight dye can be expected to stain erythrocytes, cytoplasm and muscle. The large molecular weight dye can be expected to stain collagen. The authors particularly recommended the following combinations:–
 
General use  –  AO10, DR80
After PAS    –  AY24, DY11
After silver   –  AR4, DY11

Notes

  1. This technique has not gained favour as it uses some difficult to obtain dyes.
  2. Picro-mercuric-alcohol is a saturated solution of both picric acid and mercuric chloride in absolute ethanol. It is usually applied to sections overnight after dewaxing and treating with ethanol, but before washing with water. If used, the iodine-thiosulphate sequence should be used to remove mercury pigment before staining.

 

Reference
Lendrum, A.C., Slidders, W. and Fraser, S., (1972),
Renal hyalin: A study of amyloidosis and diabetic fibrinous vasculosis with new staining methods.,
Journal of Clinical Pathology, v 25, page 373

 


 
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Last updated May 2005.