[Histonet] Frozen sections and loss of nuclei?
MinHan.Tan <@t> vai.org
Wed Dec 10 23:47:59 CST 2003
Hi there Histonetters,
I would like to seek your advice about a problem I am having with frozen
I am working on some frozen tissue dating back to about 1998, and
performing IHC on parathyroid tissue with p27 antibody. I use the
I have noted that certain samples (and particular samples only) exhibit
loss of nuclei in the central region (an amorphous blob of tissue
extending to near the margins, about 70-80% of the specimen) (no
staining with hematoxylin either). Any nuclei that stain with my IHC in
that area are very faint, and there is no hematoxylin counterstained
nuclei in the entire region. Nuclear staining seems to be fine near the
margins - of course it can be argued that these are artifacts. Where
there are folds of tissue, both nuclear and background staining is
prominent. Other samples of the same type of tissue seem to stain OK
throughout the tissue. No problems with paraffin-embedded tissue at all,
the nucleus stains fine in every sample I have tried.
It is unlikely to be due to underfixation, since I immerse my 5 micron
unfixed slide in neutral buffered formalin for 30 min prior to IHC. I
see it regardless of whether I perform HIER on my frozen sections or
not. (yes, I do perform HIER on my frozen sections)
I wonder if the following explanations are plausible:
(a) tissue has degraded - but it would not make sense for degradation to
take place from inside out.
(b) sections may be too thin for some reason? (mine are 5 micron) - but
this might explain why only the borders and folds stain.
Van Andel Research Institute
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