[Histonet] The unjoys of working with cryo rat turbinates
gcallis <@t> montana.edu
Thu May 6 09:48:49 CDT 2004
This is the reason we invested in Cryojane for the majority of
undecalcified or decalcified FS and difficult soft tissue FS work. If you
want to do antigen retrieval on any kind of frozen section of bone,
calcified or decalcified or even soft tissues, they will love to fall off
the plus charge slides and annoy you into tears or fits of bad language.
I do have one fellow who decalcifies in EDTA on
periodate/lysine/paraformaldehyde (PLP) perfused, then immersion fixed
mouse turbinates at 4C, with one change of this fixative. He cryoprotects
during decalcification with 10 % sucrose/1.25% EDTA, tetrasodium in PBS
until head is done (1 to 2 weeks, rat will take longer than mouse) snap
freezes and mounts his FS on plus charge slides. We can H&E on these but
the rinsing is minimal/gentle not with running water. We also use Richard
Allan progressive Hematoxylin 1, do not clarify, blue, rinse briefly in tap
water, go to 70%, eosin, etc, etc.
However, for IHC, he NEVER does retrieval but if you antigen is compromised
by cross linking, you may be better off with fixation then EDTA
decalcification - NO frozen sections at all IF you need to do retrieval.
Processing rat turbinates needs to be done on an extended processing
schedule - if you need one, let me know, it is easy to set up.
Discouraging but possible, tests your patience,creativity, maybe the
pocketbook if you buy Cryojane.
I'm off to undecalcified mouse snout FS this A.M. but Cryojane is assisting
Research Histopathology Supervisor
Veterinary Molecular Biology
Montana State University - Bozeman
PO Box 173610
Bozeman MT 59717-3610
406 994-6367 (lab with voice mail)
406 994-4303 (FAX)
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