mice frozen tissue IHC protocol.

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From:"Masayuki Miyagishima, MD" <mmiyagis+@pitt.edu>
To:HistoNet <HistoNet@Pathology.swmed.edu>
Date:Sat, 27 Mar 1999 23:36:11 -0500
Content-Type:text/plain; charset=iso-2022-jp

This is a very fundamental question.
What is the best or standard IHC protocol for FROZEN MICE heart and
They are 4% paraformaldehyde perfusion fixed and sit for 2 hours in ice
cold 4 % paraformaldehyde and then treated with 30 % sucrose for
overnight, then embedded in OCT and frozen in Isopentan.  The tissue is
limited and small, but we need to do many IHCs.

1) When we cut the frozen tissue samples, we cut about 50 sections at
once and put them into #027#(IP#027#(J70 fridge for the future IHCs.  Do you think I
should refix them and air-dried before putting them into #027#(IP#027#(J70?

2) After taking out from #027#(IP#027#(J70 fridge, we treat the sections with 10% Goat
serum, then treat with rabbit primary antibody for 2 hours at RT, then
with goat anti-rabbit secondary antibody for 2 hours at RT.  Do you have
any modifications for mice tissue?

Thank you in advance.

Masayuki Miyagishima, MD
University of Pittsburgh, 
Department of Surgery
412-647-2345, and ask the hospital operator to page 3228, please.

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