In situ hybridization
K J C 944
kjc944 at aol.com
Wed Jul 10 09:39:45 EST 1996
I'm getting ready to try whole mount in situ hybridization on mussel
larvae. These larvae are, for the most part, less than 200 microns in
diameter. While I understand the hybridization process, I am confused
about sample preparation. I plan to fix the larvae in 4%
paraformaldehyde, 1X MOPS buffer. My question is, how long do I fix my
larvae? I know that fixation time is critical, but I have seen protocols
in which the fixation time is anywhere from 30 minutes to 48 hours. I
don't have alot of larvae to mess around with and was wondering if there
is a rule of thumb for fixation time, depending on the size of the sample.
My second question has to do with
dehydration. Since I will not be imbedding my samples in parafin for
sectioning, is it still necessary to dehydrate? Couldn't I just fix,
quench, and store in buffer at 4C until I'm ready to hybridize?
Thanks in advance!
College of Marine Studies
University of DE
Lewes, DE 19958
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