[Histonet] Masson Trichrome

From:"Hermina Borgerink"


Below is a modified version of the original Masson's Trichrome which I
have successfully used for over 40 years.  Instead of using
Woodstain-Scarlet, which I don't think is available anymore, substitute
Crocein Scarlet which Sigma sells. 

Masson's Trichrome (Modified)    

Post-fix in Bouin's fixative overnight if primary fixation was not done
using Bouin's

1. Bring sections to water and wash for 5 - 10 minutes.
2. Stain nuclei for 15 minutes in Weigert's iron hematoxylin
3. Wash for 5 - 10 minutes in running water and rinse in 1% acetic acid.
4. Place slides in Woodstain Scarlet-Acid Fuchsin mixture for 5 minutes.
5. Rinse briefly in 1% acetic acid and differentiate in 5%
phosphomolybdic acid.  Differentiate to the point where collagen and
ground substances are pale pink to colorless, usually about 30 seconds
to 2 minutes. 6. Rinse in 1% acetic acid. 7. Place for 15 - 20 seconds
in the Aniline Blue solution (Aniline Blue - Woodstain Scarlet - Acid
Fuchsin mixture). 8. Place for a quick rinse in 1% acetic acid. 9.
Dehydrate in four changes of alcohol. 10. Clear in xylene and mount in

Preparation of Staining Solutions:

Weigert's Hematoxylin:
Solution A:
29% ferric chloride - 4 ml
Distilled water     - 95 ml
Conc. HCl           - 1 ml
Solution B:
Hematoxylin - 1 gm
95% ethanol - 100 ml
For use, mix equal parts of solutions A and B.  Use fresh each time

Woodstain Scarlet-Acid Fuchsin:  prepare a 1% solution of Woodstain
Scarlet NS and a 1% solution of Acid Fuchsin (C.I. no 692).  Mix 8 parts
of Woodstain Scarlet solution with 2 parts of Acid Fuchsin solution and
add 1ml acetic acid to 99ml of the mixed dye solution.  

Aniline blue:  To 85ml of 1% acetic acid add 5ml of a saturated solution
of Aniline Blue (C.I. no 707) and 10ml of the above Woodstain
Scarlet-Acid Fuchsin mixture.

Hermina M. Borgerink, BA, HTL(ASCP)QIHC
Wake Forest University Health Sciences
Department of Pathology
Medical Center Blvd.
Winston-Salem, NC 27157
Tel. (336) 716-1538
Fax (336) 716-1515
e-mail hborgeri@wfubmc.edu

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