[Histonet] CD40

[Reynolds,Donna M]

What are you using for Endogenous blocking? If you are using methanol with H2O2 that may be your problem. Many CD antibodies are very sensitive to the methanol. Try PBS or whatever buffer you are using to dilute the H202. I would also try going straight form the -20 into cold acetone without air drying and fix for10 min. 

Donna Reynolds HT (ASCP)
U. T. M.D. Anderson Cancer Center
Chief histology Tech, Department Cancer Biology
713-792-8106
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Message: 3
Date: Mon, 9 Jul 2012 07:55:16 -0700
From: Bea DeBrosse-Serra <BDeBrosse-Serra <@t> isisph.com>
Subject: [Histonet] Need help with CD40 on frozen sections
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Histonetters,

I need some help with CD40 on frozen sections (spleen, non-treated). I cut my frozens, let them briefly air dry and keep them at -20?C. When I use them for staining, I air dry them, fix them in cold acetone for 5 minutes and start the staining process. I use the primary AB from BD Pharmingen, # 550285 at various dilution, incubate them overnight at 4?C, use the secondary HRP conjugated antibody, DAB, counterstain and don't get any staining. Does anyone have any pointers, suggestions?

Thank you in advance,

Bea

Beatrice DeBrosse-Serra HT(ASCP)QIHC
Isis Pharmaceuticals
Antisense Drug Discovery
2855 Gazelle Ct.
Carlsbad, CA 92010
760-603-2371
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