immunocytochemistry - 2nd Ab problems

Barb McCracken bmccrack at students.uius.edu
Wed Sep 4 07:30:27 EST 1996


I am trying to do immunocytochemistry on some Bouin's fixed intestinal
tissues.  I am having trouble with staining in my negative control
tissue.  It is not just high background but specifically stained cells. 
My primary is ascites made in a mouse against a pig antigen.  For my
second antibody I am using a biotinylated goat anti-mouse IgG, heavy and
light chain specific, Fab fragment only.  I am using the Histomark Red
protocol from KPL.  To control background I am using goat serum and a soak
in buffer with BSA.  These measures have significantly reduced the
background but I have beautifully stained, obviously positively stained,
cells in the tissues without promary antibody.  Does anyone have a
suggestion?

Thanks,
Barb



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