Paquin & Goddard's Trichrome
for Elastic and Collagen

Solutions

Paquin & Goddard's iron hematoxylin or equivalent

Solution A
Eosin Y	0.7	g
Phloxine	0.3	g
Orange G	0.1	g
Phosphotungstic acid	1	g
Distilled water	1	L

Solution B
Phosphotungstic acid	2	g
Distilled water	1	L

Solution C
Acetic acid, glacial	4	mL
Distilled water	1	L

Solution D
Aniline blue	0.4	g
Acetic acid, glacial	10	mL
Distilled water	990	mL

Tissue sample
5µ paraffin sections of Masson's picric-chrome-formalin fixation was specified. Most trichrome stains benefit from picric acid or mercuric chloride fixation. The specified fixation may be compensated for by secondary fixation of sections in Bouin's fluid.

Method

1. Bring sections to water via xylene and ethanol.
2. Stain with Paquin & Goddard's iron hematoxylin.
3. Wash with water.
4. Place into solution A for 5 minutes.
5. Place into solution B for 5 minutes.
6. Rinse twice with solution C.
7. Place into solution D for 5 minutes.
8. Rinse twice with solution C.
9. Place into solution B for 5 minutes.
10. Place into solution C for 30 minutes.
11. Rinse quickly with 95% ethanol.
12. Dehydrate with iso-amyl alcohol.
13. Clear with toluene and mount with a resinous medium.

Expected results

• Nuclei  –  black
• Elastic  –  red
• Cytoplasm  –  pink
• Collagen  –  blue

Notes

1. If iso-amyl alcohol is not available, t-butanol is often suitable when the staining is likely to be removed with ethanol, although rapid dehydration with ethanol can often overcome this.
2. Although toluene was specified for clearing, xylene is likely suitable.
3. Masson's picric-chrome-formalin contains:
    
   

   Picric acid, sat. aqu.	187.5	mL		Add the alum to the formalin and leave for 1 hour.
   Chromium aluminum sulphate	7.5	g		Add the picric acid solution.
   Formalin, Conc. 40%	67.5	mL		Let stand for 24 hours, and filter.
   Return to fixation.

 

Reference

Gray, Peter. (1954)
The Microtomist's Formulary and Guide.
Originally published by:– The Blakiston Co.
Republished by:– Robert E. Krieger Publishing Co.
  Citing:–
    Paquin & Goddard, (1947)
    Bulletin of the International Association of Medical Museums and
    Journal of Technical Methods, v.27, pp.198

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