On Mon, 24 Jan 2000, Gayle Callis wrote:

> I believe the importance of the Bouins, with the acetic and picric acid
> components, is to acidify the tissues, in order to make the dyes penetrate
> correctly.  John Kiernan will help on this.

    I don't know; had imagined it was a post-fixation. This
    step isn't needed if the primary fixation is Bouin, Zenker
    or Susa. For sections of formalin-fixed objects, you can use
    just a picric acid solution (without formaldehyde & acetic
    acid) and get the same improvement in Masson staining.

> The publication in J of Histotechnology, Citrate buffer alternative to
> picric acid for Masson trichrome stain, v 10 (4), Dec 1996 by Joyce Moore
> gives details.
>
> It is a 0.1M citrate buffer, pH 5.9 to 6.0, microwaved for 1 min. I would
> think the important factor in this solution is that the pH of the buffer
> matches that of Bouins, would be a good thing to check, and try.

    Does this paper say if pH 5.9-6.0 was the best of several
    tried? If only one pH was tried, it might be the heat alone
    that is the effective pre-staining treatment. In any case
    it cannot be just a pH effect because Bouin's fluid
    is much more acidic: pH 1.5. A saturated aqueous
    solution of picric acid has about the same pH. A picric acid
    solution alone causes shrinkage of liver to 74% of original
    volume, but in Bouin's this is compensated for by the acetic
    acid, which would cause swelling if used alone. (Info from
    Baker's "Principles," Ch. 5 & 6)

    One could speculate about how acids, picrate ions, citrate
    ions etc might work, but theories would be difficult to
    test without a lot of experiments. I can't see a funding
    agency giving anyone a grant to do them!

 John A. Kiernan,
 Department of Anatomy & Cell Biology,
 The University of Western Ontario,
 LONDON,  Canada  N6A 5C1