in situ/immuno combo

David M. Berman eyeball at netcom.com
Thu Apr 7 09:03:34 EST 1994


KGEISS%RAVEN at EAGLE.AAMRL.WPAFB.AF.MIL wrote:
: Has anyone out there done both in situ hybridazation (for RNA) and immuno 
: (antibody) detection on the same slide?  In our in situ protocol, we digest the
: tissue with proteinase K, it seem to me that this would affect our ability to
: be able to detect the related protein at a later time.  And what if we did the
: protein detection first before the in situ, would this work.  I don't know what 
: detection methods would be suitable to this regimen.  Any insight would be
: greatly appreciated.

: Thanks,
: GEISS...[:)

I haven't done it but there's a protocol in "Current Protocols in
Molecular Biology" (the red book)  I HAVE done immunohistochemistry
after proteinase treatment (pronase) which also works well for in situ
on the same tissues.  For immunohisto., some antigens are better
detected after protease treatment, some go away.  My advice is to
optimize protease treatment carefully for each procedure and then
combine them.  If your Immuno. procedure doesn't tolerate protease,
try tritonX-100 and/or the microwave/citrate procedure (ask Vector
Labs for this protocol if you need it).Good luck.
-- 
                                             eyeball at netcom.com



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