I've tried absorbing antibody with tissue to some success. I your case it would be lymph node homogenate using protease inhibitors. Absorb 2-3x, spin out the debri and titer appropriately. -----Original Message----- From: Ronald Rabin [mailto:RRABIN@niaid.nih.gov] Sent: Monday, September 25, 2000 5:00 PM To: cyto-inbox Subject: high bg fluoresc micrscpy I am staining human secondary lymphoid tissues (frozen sections) with goat and rabbit polyclonal antibodies. The secondary antibodies are rabbit and goat HRP respectively. I use the Vector ABC system to amplify. I am having huge problems with background non-specific staining. I have tried airfuge prep of antibodies, blocking with the serum from the same species as the secondary antibody. It is not the ABC system per se, or even the secondary antibody, as signal is low without primary controls. Anybody know how to block nonspecific binding? Ronald L. Rabin, M.D. Research Fellow Cytokine Biology Unit Laboratory of Clinical Investigation NIAID/NIH Building 10/Room 11N228 10 Center Drive MSC 1888 Bethesda, MD 20892-1888 Phone: 301-402-4910 FAX: 301-402-0627
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