Robert, We routinely fix human whole blood samples with 1% formalin (diluted from 10% stock methanol free from Polysciences) in 10mM Hepes/saline prior to labeling with antibodies specific for P-selectin. While I can't speak for mice, at least in humans we do not see any increase in P-selectin expression in unactivated samples using this "fix-first" technique. Lori Krueger Center for Platelet Function Studies University of Massachusetts Medical School ----- Original Message ----- From: "Jensen, Robert" <robert-j-jensen@uiowa.edu> To: cyto-inbox Sent: Friday, November 09, 2001 9:33 AM Subject: Murine Platelets. > > We have been doing some experiments looking at markers on activated mouse > platelets. Now a member of our lab wants to do a time course of expression > following thrombin induced activation. The plan was to remove samples at > various time points following activation, fix, and then stain all sample at > the same time at the end of the time course. But we have noticed that > non-thrombin treated platelets appear positve for P-selectin when we fix > with 1% paraformaldehyde (unfixed are negative so we are not activating > during the isolation procedure). > > Does anyone have a fixation method they use prior to staining platelets that > does not cause them to up regulate P-selectin? > > thanks > Robert Jensen
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