IHC on frozen cryosectioned rat kidney

From:"Stone, Jessica" <JStone@ligand.com>


If you have time and are so inclined, can you write me the best (or
successful methods in your hands) for initial freezing of tissue (rat kidney
~5mm thick); and post cryo-section processing for immunohistochemistry
(doing indirect with Cy3).  I am a beginner and having problems with cell
damage and inconsistent results; I believe culprit is initial freezing
method or tissue treatment after sectioning i.e. dry or not to dry sections,
if dry then before and/or after fixation, best fixative and time in
fixative, does the tissue need to be perfused etc. etc.

Thank You for your time and help!!

P.S. any thoughts on switching to a  fixed, paraffin embedded method?? I
tried it once and got no signal, not sure if overfixed, overprocessed, or
Cy3 bad choice...

Jessica Stone
Ass. Scientist
Ligand Pharmaceuticals
La Jolla, CA

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