Re: Trichrome stains.

From:Lee & Peggy Wenk <> (by way of histonet)

Peggy Wenk here.

Quality Assurance specialist for Masson Trichrome
and MSB, huh?

Well, I'll give a try at this one. Differences
that people do in the two stains.

I'm assuming that the Brancroft/Stevens book
would be the standard in England. So I'll talk
variations from that.


1. Like using a Ferrous hematein stain, instead of
Weigert Hematoxylin.

Can be reused for about 2 months, and the nuclei are black
after staining with the red and blue dyes.

2. I like using the phosphotungstic acid, instead of
phosphomolybdic acid.

3. I prefer either a combination of acid fuchsin and
ponceau de xylidene, or just ponceau S, instead of
acid fuchsin.

Get a redder red.

4. We use aniline blue instead of methyl blue. Different
shade of blue. Our pathologists like this shade.

5. Brancroft/Stevens version does not mention post-mordanting
with picric acid if originally fixed with NBF.

It does mention removing the mercury ppt., so I'm assuming that
they  say this stain should be done on mercury fixed tissue.
Since most of ours is formalin fixed, we usually post-mordant
with the picric acid Bouin solution at 60 degree C. for
45-60 minutes.



1. There is no mention of post-mordanting if formalin fixed.
We used to use the Bouin solution at 60 degree C. for 45-60 minutes,
but now we use a solution of 30 mL 3% potassium dichromate in water
plus 10 mL 10% HCl in 95% alcohol. Mix together just before use,
and postmordant at room temperature 5 minutes, then wash slides
in running water for 1-5 minutes, until color disappears.

This was something one of my students adapted from some other
procedure, and we like the time savings. Plus we got rid of
picric acid. However, there are dangers with potassium dichromate.

(We tried it with the Masson trichrome, but it was splotchy
in the final color results.)

2. Nuclear stain - the original procedure calls for a double
nuclear stain of Mayer hematoxylin and Celestine blue.

We switched to the Weigert hematoxylin for the nuclear stain,
and have since switched to the Ferrous hematein.

We like the black nuclei better.

3. Instead of brilliant crystal scarlet, I have used the
red dyes listed above in the Masson trichrome, with good

However, since we sent all the way to England to
get the brilliant crystal scarlet, and still have about
80 grams left out of the original 100 grams from 20 years
ago, we'll keep using it until it's gone.

4. Instead of methyl blue, we use the aniline blue from
the Masson trichrome. Our pathologists are used to that
shade of blue and like it better.

Hope this is what you were looking for.

Peggy A. Wenk, HTL(ASCP)
William Beaumont Hospital
Royal Oak, MI 48073 wrote:
> Dear histonetters,
> Having recently been appointed as MSB/Massons Trichrome specialist for our
> External Quality Assurance scheme I would be grateful if anyone could give me
> information on any non-standard techniques that work well or any tips on
> performing the perfect trichrome stain (MSB in particular). Many thanks.
> Jackie. West Midlands. England

<< Previous Message | Next Message >>