[Histonet] Help With Cutting Heart


Hello Histonetters,
   I have been cutting transverse sections of rat heart using the following protocol:
1) Dissect heart, cut in 3mm disks
2) Fix overnight 4% PFA @ 4 deg.
3) Cryoprotect in 30% sucrose around 5 days.
4) Embed in OCT and freeze blocks in isopentane that has been
   cooled in liqid N2 for 1min.
5) Store 6 hours at -80 deg.
6) Store o.n. at -20 deg and cut 16uM on cryostat also at this
   The quality of the resulting histology is o.k but not great, specifically the endocardium is a bit torn.   I cannot parraffin embed because of downstream in-situ hybridization signal intensity problems so i must use a frozen or perhaps fresh tissue histology protocol. Does anyone have any suggestions or experience with good frozen heart histology, any changes i could make to my protocol? maybe a new protocol? Please help! 
Thanks in advance,
Fred Grau
Dept. Neurbiology & Behavior
SUNY Stony Brook

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