cytokine staining

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From:Gayle Callis <>
Date:Thu, 22 Jul 1999 10:11:37 -0600

Pharmingens haws monoclonals for cytokines, but the immunostaining is difficult.
Labs who have had the most success use these clones, and it it well
documented in the literature.

The literature, for murine cytokine IHC, is done on frozen sections, with
some groups successful with formalin fixation on frozens, using a saponin
permeabilization of cell membranes.  A couple of labs have used acetone
fixed frozen sections, one with a saponin method, another with TRIS buffered
saline containing Tween 20.   

To date, Pharmigen has done only immunocytochemistry on cells stimulated
to produce cytokines, and have a very nice protocol, BUT tissue sections
have not been worked out by them, in other words, you will be on your own
attempting to make the intracellular immunolocalization of IL-6 work.

I strongly recommend using monoclonals, and very good secondaries, some
labs have used F(ab')2 secondaries, absorbed to mouse.  IF you succeed
with your cytokine IHC, there are people waiting in the wings to know
what you have done. Be sure you use DAB, enhanced is nice to have the
most sensitivy.

I do not know anyone using paraffin on the IL's for mouse, I think there
has been some success with TNF on NBF/paraffin, but frozens seem to be norm.

Have been working on cytokine staining steadily for some
months and still have poor success, will continue to tweak until the cows
come home and the cytokines stain!

Good luck, you will truly have your work cut out for you.

Gayle Callis

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