Staining problem details

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From:Pam Plumlee <> (by way of histonet)
To:histonet <>
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Hi: To add more details to the staining problem I'm having...first, the
hematoxylin staining looks normal..just no pink at all in the cytoplasm.
I move the rack of slides when they are melting off the paraffin in the
oven, so when staining there is more than enough solution in the
staining dishes.  I filter the hematox. often and change and rotate the
alcohols and xylene.  My routine H&E protocol is just about the same as
in Carson's Histotechnology book.  I have added a little time to
deparaffination and use D.I. rinses before putting the slides in the
Eosin Y Alcoholic because our tap H2O is very acid.  The puzzling part
of this for me is the staining can be uneven on the same section,
microscopically the section does not look thick or thin and there does
not seem to be any paraffin left on the unstained section.  My slides
are not dried in the microwave.  This may just be a ph problem or
perhaps a processing problem or just a lab gremlin problem.  thanks for
the suggestions so far. pam

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