sectioning of frozen tissues
Greetings histology experts,
We are doing immunohistochemistry on frozen sections but so many
times the morphology of the sections (especially small tissues such as
mouse thymus) are terrible. Consequently we've been looking for
alternative protocols. We are considering the protocol listed on the
Pharmigen web site. Part of the protocol is as follows:
II. Sectioning of Frozen Tissues
1.Before cutting sections, allow the temperature of the block
to equilibrate to the temperature of the cryostat (-20#176#C).
2.Place the tissue block on the cryostat specimen disk. Adjust
the positioning of the block to align the block with the knife blade.
Cut tissue block until the desired tissue is exposed.
3.Cut sections of the desired thickness (usually 5 um), place
the sections on a Fisher Superfrost slide and dry overnight at room
4.Fix slides by immersion in cold acetone (-20#176#C) for 2
minutes or other suitable fixative (e.g. alcohol, formal alcohol,
formalin, etc.), air dry at room temperature and proceed to staining
5.Alternatively, the frozen section slides can be stored for a
short period of time at -70#176#C in a sealed slide box. When ready to
slides from freezer and warm to -20#176#C in the cryostat or
-20#176# freezer, fix for 2 minutes in cold fixative (acetone or other
and allow to come to room temperature to continue with the
Now, my question is wouldn't allowing the section to dry overnight
at room temperature (as described in step 3) result in destruction of
certain proteins (maybe ones we want to stain for)?
Thank you in advance,
Washington University School of Medicine
St. Louis, MO
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