----- Original Message -----
From: J. A. Kiernan <jkiernan@julian.uwo.ca>
To: Bryan Llewellyn <bryand@netbistro.com>; Histonet
<histonet@pathology.swmed.edu>
Sent: September 13, 1999 10:39 PM
Subject: Re: Mast cells/Aldehyde-toluidine blue


>
>   Dear Bryan,
>
>   This is one I haven't come across before. Have you got a
>   reference for it, or is it a Llewellyn original? Also, what colour
>   are the mast cells, blue or metachromatic? Is it an unremovable
>   stain like aldehyde-fuchsine, which resists acid-alcohol - so that
>   you can do it first and then follow up with other methods that
>   would remove an ordinary basic dye? Lotsa questions. No hurry,
>   but I'm interested because mast cells in skin & lung, and also in
>   the brains of a few species (notably the hedgehog) were my
>   major source of research grant money in the 1970s and '80s.

I hesitate to call it an original, but I did develope it.  I never pursued
publishing it because of the variability with the toluidine blue.
Incidentally, other thiazine dyes work too, just not as well.  The only
toluidine blue I could find that is consistent is the one Fisher sells.
There must be a reason for this, but what it is I haven't a clue.  Perhaps
chromatographic analysis of various dyes would help determine why.  The data
published years ago on which the method is based is commented on in
Culling's second edition, (page 214, 250 in 3rd ed.), where a list of dyes
is given that can be used in Schiff type reagents, i.e. dyes that can be
linked with aldehydes.

Mast cell granules are stained very dark blue, not metachromatically.  I
presume the dye remains attached similarly as basic fuchsin does by some
kind of ionic bond, but I have not tested it.  Incidentally, there is only
very poor elastic staining.  Basic fuchsin is known to sometimes attach via
dipole interactions, but toluidine blue does not appear to do so as easily,
if at all.  Due to that, I presume, the background is essentially clear.
What mucin staining there is pesent, is pale blue and insignificant.  The
mast cells are very visible in high contrast.  I counterstain with nuclear
fast red-tartrazine.  Tartrazine is a straightforward acid dye, many of
which often remove basic dyes during staining.  This does not.


>   Perhaps Ald-Tol-Blu is in your StainsFile web page, and I've not
>   spotted it yet. The "pick a letter" method of looking things up
>   doesn't let you browse easily, as through a contents list or
>   an index.

It isn't on there.  In fact, I did very little for over a year because of
health concerns.

Do you think I should have a single pick list of methods?  I have spent
quite some time trying to figure out an easy to use interface.  Currently it
is a mix of alphabet selection and preselected method types (hemalums,
trichromes), but I am not really satisfied with it.

Suggestions from others on histonet would be most welcome.


All the best,
Bryan