Re: [Histonet] cartilage staining

From:Rene J Buesa (by way of histonet)

Dear Beppe (and other):
According with Bancroft & Stevens (1977) when discussing the Standard
Toluidine Blue (S.T.B.) method by Wolman (1971) they attribute the cartilage
methachromatic staining  to "probably" an electrochemical bonding that,
with Cooper (1974) is minimised if the staining solution is saturated with
The electrochemical bonding has to occur between the sulfur in the toluidin
blue molecule and the chondroitin sulfates molecules found in cartilage.
Just an additional detail!
Rene J.

John Kiernan  wrote:
Dear Beppe (and all others),

I think the reason toluidine blue is a more pleasing
cartilage stain than safranine O has to do with the shape
of the dye molecule. Toluidine blue cations are flat,
and can pile up in stacks of two or more, so that each
half-sulfate-ester anion in the cartilage matrix can
bind more than one toluidine blue cation, resulting in
intense staining. The stacking also accounts for a
shift in the wavelength of maximum absorption. This
results in the pleasing metachromatic red-purple colour
of cartilage, mast cell granules and some types of mucus,
while cell nuclei (DNA) and basophilic cytoplasms (RNA)
are blue.

Safranine O (which is always a mixture of isomers,
because of the way it's made) has much more bulky
cations than toluidine blue. They may form aggregates
but they cannot stack up tidily. The dye will (like any
other cationic dye) stain cartilage fairly selectively
if applied from a solution too acidic to stain DNA (eg
at pH below 2). If you don't need a counterstain, that's

John Kiernan
Anatomy, UWO
London, Canada.
Rene J Buesa wrote:
> Toluidine blue has always worked better for me. Both have a similar
>chemical composition
> which for Safranin-O is C20 H19 N4 Cl (MW=351) and for Toluidene Blue is
>C15 H16 N3 Cl and S (MW=238).
> Their difference is not in the fact that Toluidine Blue has a smaller
>mollecular weight, but
> because Toluidine Blue has Sulfur in its mollecule. Probably this is the
>reason for
> a better affinity with the Sulfur present in cartilage.
> Rene J.
> Beppe Intini wrote:
> Dear Histo-Friends,
> I hope you can help me again....!
> Just a quick question: is Safranin-O better than Toluidine blue for
> cartilage staining? And Why?
> Thanks
> Beppe Intini
> --
> Giuseppe Intini
> Clinical Assistant Professor - Periodontics
> PhD Candidate - Oral Biology
> University at Buffalo
> Buffalo, NY 14214
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