Hello, I have a problem with a cell surface antigen that is (I believe) internalized. I have an antibody which binds well. And I visualize with a fluorescent secondary. I can show that over time, the signal decreases, I believe, due to internalization. But I would like to show that the missing signal is from primary antibody actually inside the cell, and therefore sequestered from the secondary, and not from the primary just becoming unbound over time. I have tried to do this by permeabilizing the cells after treatment with primary and internalization. But after permeabilization, I lose all signal. (Probably due to the primary being washed away?). My question is this. Does anyone know of a good procedure I can use to permeabilize mammalian cells after treatment with antibody that will not destroy my signal due to washing away or even destruction of the secondary/primary interaction? Thanks. Mark D'Amico, Ph.D. Scientist Panacea Pharmaceuticals 207 Perry Parkway #2 Gaithersburg, MD 20877 (240) 243-8000 Ext. 206 mdamico@panaceapharma.com There is a school of thought that all of the universe is nothing more than a figment of my own imagination. But in that case, where the hell are all the moon maidens?Received on Fri Feb 17 14:58:00 2006
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