Dear Histonetters,

We have been doing some cryosectioning of murine lenses for
immunohistochemistry, and have been having much difficulty in getting good
looking sections.  Of particular problem is the shattering of the lens,
especially towards the center.

We have attempted a variety of different methods, including frozen sections
that are post-fixed with methanol and acetone and previously fixing the lens
in 4% paraformaldehyde for an hour.

Are there any other kinds of fixative that you may recommend?  I have heard
of Davidson's fixative and was wondering whether this would disrupt the
actual IHC stainings.

We have also tried 4% fixation, and while the lens itself looks good when
initially cut, giant cracks begin to form over a short period of time while
the slides have been in the cryostat.  I have also tried leaving the slides
out at room temperature after doing sections to see if the shattering is
reduced, but the staining was completely gone, I think possibly because the
cytoplasm may have leaked out as a result.

Any kind of recommendation, techniques, and so forth are definitely
welcome.  Thanks.

David Park
szero1009@gmail.com
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