[Histonet] Cryotomy help

[Carl Hobbs]

I assume that the fresh tissue was fixed in methanol?
If so, it is irreversibly coagulant- fixed ( like boiling an egg white...no going back)
Place the specimen in dist water until it melts to RT
x3 changes of dw 1hr each at RT on a gentle rocker to get rid of methanol ( antifreeze, hence inability to freeze as you have replaced water)
Place in OCT at RT on a rocker for 30 mins
Orientate and snap-freeze
Or, if your mass spec doesn't like OCT....freeze without OCT
?
NB: Most important to freeze quickly to avoid ice-crystal formation
Proper snap-freezing is used to make the water go from liquid to vitreous ( glass-like) state, avoiding the ice-crystal state ( which will occur with SLOW freezing....thus giving you horrible ice-crystal artefact)
Good luck!

Carl Hobbs FIBMS
Histology and Imaging Manager
Wolfson SPaRC
Guys Campus, London Bridge
Kings College London
London
SE1 1UL

020 7848 6810