AW: [Histonet] Proof that fixation in formaldehyde solution can bereversible?

[Gudrun Lang]

 Sorry Rene,
but isn't it the other way round? That penetration is rather fast and
fixation in the sense of making methylenbridges is rather slowly? So tissue
of 5 mm is penetrated within a few hours, but fixation takes much longer?

I also learned that fixation with formaldehyd can be reversed by washing in
tapwater for several weeks. In some old books they recommend washing the
tissue the same duration as fixation was done.

Gudrun

-----Ursprüngliche Nachricht-----
Von: histonet-bounces <@t> lists.utsouthwestern.edu
[mailto:histonet-bounces <@t> lists.utsouthwestern.edu] Im Auftrag von Rene J
Buesa
Gesendet: Dienstag, 03. Oktober 2006 19:05
An: GT Hebert; histonet <@t> lists.utsouthwestern.edu
Betreff: Re: [Histonet] Proof that fixation in formaldehyde solution can
bereversible?

GTH:
  A piece of tissue sitting in PBS for 1 month after being fixed for 24-48
hours only is likely to have suffered tissue decomposition at the center of
it, specially if it was a thick piece. Formalin is an extremely fast
fixative but with a very slow rate of penetration, this would have
determined external fixation but not complete fixation.
  Fixation is not really reversible in a strict way and that is why you need
a strong Heat Inducet Epitope Retrieval (HIER) before IHC to "undo" or
"unfix" the crosslinkage produced by formalin.
  The damage to the tissue could have been caused by improper storage = PBS
at room temperature during 1 month. If processed that tissue is very likely
to present altered microscopic appearance.
  Just my opinion!
  René J. 

GT Hebert <emerald_lake77 <@t> yahoo.com> wrote:
  Hello,

I am in a heated debate with an investigator regarding his samples. They
were shipped to me after (24-48hrs fixation - 10% NBF or 4% PF unsure
exactly which one) and then switch to PBS for shipment. They sat at room
temperature for over 1 month before being processed routinely and embedded
in paraffin wax. 

Can someone tell me if indeed the fixation is reversible, or once they have
been fixed for over 24hrs they remain fixed?? Can anyone refer me to books
or papers that talk about this?? 

Also, what affect on antigenicity will such storage in PBS have on these
samples?

Thank you all so much for your help.

G. H.
Cambridge, MA



Key words:
NBF, Alcohol, 10%, paraformaldehyde, 4%, fixation, fix, ethanol, 70%,
reverse, reversible.


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