IHC on mouse or rat tissue & CSA related problem

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From:Ms Louise Taylor <179LOU@chiron.wits.ac.za> (by way of histonet)
To:histonet <histonet@magicnet.net>
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Hi Histonetters

I have 2 queries ? I have always done immunohistochemistry (IHC) on
human tissue. I understand the theory and enjoy doing IHC . I am
however curious about the protocol when doing IHC on mouse,rabbit or
any other host . Does one need to use a host absorbed secondary
antibody  or is it type specific? I gather that there may be cross
reaction from species to species. Any information on this would be
helpful. Thanks in Advance.

Second question: Has anyone used the CSA (Dako) kit; It is great to
amplify minuuuute antigenic sites, however I do sometimes get a lot
of non-specific staining. I normally when using the CSA kit , use an
avidin/biotin block anyway. This strong punctate dot-like signal on
the membrane of cells is on both the test as well as the negative
control. Has anyone experienced the same problem? Should I dilute the
primary even more or is there something else I could try? I have
tried x-tra washings as well as Triton -X-100/BSA in my buffers.

All help is appreciated.


Zenobia Haffajee
Anatomical Pathology
South Africa

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