Re: IHC on frozen muscle

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From:Joyce Kotzuk <>,
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I had terrible trouble with fixed paraffin embedded muscle not staying on the sections also, and found that if I cut them at 4-5 microns, they stayed on  better than at my usual 8-10 microns. But unfixed frozens, I don't know...

>>> Louise Taylor <> 06/08/00 07:31AM >>>
Hi histonetters,

I am doing dystrophy markers (dystrophin, sarcoglycan etc) on cryostat
sections of  fresh frozen muscle. These biosies have been frozen in
isopentane cooled in liquid nitrogen. I have noted that there is a lot of
fat in these biosies(part of the pathology?), and that section adherence
through the staining process is poor ( non existent actually) even though I
am using silane treated slides.
 I was wondering if anyone has had a similar problem, and what
countermeasures could be taken. I though that maybe fixation in acetone
prior to IHC might improve matters, but I don't know whether this will
impact negatively on the staining, I have already optimised my panel of 12
markers  on "unfixed" muscle treated the same way, and would not like to
have to reoptimise.
 Any thoughts? I would appreciate some help here, as muscle frozens are not
quite my usual sample.

best regards
louise taylor

Research Laboratory
Department of Anatomical Pathology
South African Institute for Medical Research
South Africa

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