RE: an interesting question has come up

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From:Rob Geske <rgeske@bcm.tmc.edu>
To:Gayle Callis <uvsgc@msu.oscs.montana.edu>, histonet@pathology.swmed.edu
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Gayle,

I have read this in the 4th edition of Bancroft (Theory and Practice of
Histological Techniques) and also in Thompson (Selected Histochemical and
Histopathological Methods).The following is from Thompson:

"......It is also known that formalin can form sulfhydryl groups of cysteine
to form a methylene bridge, and that reaction is reversible.  It is also
possible that at alkaline pH, formalin could reduce the disulfide linkages
of cystine to form sulfhydryl groups and proceed to develop a methylene
bridge between these groups.  Such reactions with sulfhydryl groups are
reversible and would be expected to occur at alkaline pH rather than at
neutral or slightly acidic pH of neutral buffered 10 percent
formalin......".

The references are as follows:

1.  Bennett, H.S.: The cytochemical demonstration and measurement of
sulfhydryl groups by azo-aryl mercaptide coupling with special reference to
mercury orange.  General Cytochemical Methods, Vol. 1, Danielli, J.F, NEw
York Academic Press Inc.  pp 317-374, 1958

2.  Middlebrook, W.r. and Phillips H.: The action of formaldehyde on the
cyctine disulfide linkages in wool. I  The subdivision of the combined
cystine into two fractions differeing in their reactivity toward
formaldehyde.  Biochem. J.  36:294-302, 1942

3.  Ratner, S., Clarke, H.T., The action of formaldehyde upon cysteine, J.
Am. Chem. Soc, 59:200-206, 1937.

There is also a great paper in Histochem and Cytochem by Fox in 1985.  My
understanding that "reversible" is used in the context of a chemical
reaction  not that the tissue would be in a state as it was before the
application of the fixative.  Given that formaldehyde is a polar molecule,
and in the presence of excess water, the equilibrium of the reaction would
be shifted to the left, favoring a higher concentration of reactants than
products.


rob
-----Original Message-----
From: Gayle Callis [mailto:uvsgc@msu.oscs.montana.edu]
Sent: April 18, 2000 1:06 AM
To: histonet@pathology.swmed.edu
Subject: an interesting question has come up


Have been a private discussion with someone, who brought up the point that
fixation can reverse itself, and in the context of formalin?  but that the
fixation process is eventually irreversible.

Any comments on this, I found it interesting, and know the other party is
looking in.

I have never experienced this, or even heard of it happening, but am always
ready for enlightenment, could be something I have missed in reading,
listening or discussion.




Gayle Callis
Veterinary Molecular Biology
Montana State University
Bozeman MT 59717-3610
406 994-4705
406 994-4303





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