Enclosed a reply by one of our scientists, of course I can not resist the temptation to inform you that the antibodies which are used are original CLB clones which are commercially available. John Bill, We have received your mail about platelet fixation. We are using FACS-analyses also to measure the expression of several antigens (also P-selectin). To isolate platelets from fixated blood is described in 1990 by R. Fijnheer et al (Transfusion 30: 20-25, 1990). One volume of blood was collected into nine volumes of 1% PFA (diluted in PBS). To make sure that there will be not too much activation due to the collection a large needle can be used and the first milliliters of blood can be thrown away. Make sure that during the collection of blood not too much pressure is forced (flawless collection). Blood was stored at room temperature. After one day (we have no experience with longer storage) blood was centrifuged by a soft spin and the PRP was collected. The isolated platelets could than be used for FACS-analysis. I hope that this information will be satisfying, if you have further questions you can inform us. Yours sincerely, Christine Kramer Technician of Blood Transfusion Technology CLB, Amsterdam On behalf: Dr. D. de Korte Head of laboratory of Blood Transfusion Technology John Voorn CLB Reagents Productmanager Plesmanlaan 125 j_voorn@clb.nl <mailto:j_voorn@clb.nl> 1066CX AMSTERDAM the Netherlands tel +31-(0)20 5123246 fax +31-(0)20 5123570 web site www.clb.nl <www.clb.nl> -----Original Message----- From: William Ross [SMTP:wgr8w@virginia.edu] Sent: vrijdag 31 maart 2000 16:57 To: Cytometry Mailing List Subject: platelet fixation does anyone no of a good platelet fixation procedure for the study of selectin expression? It is a study of myocardial infarction patients and timing the heart attacks is difficult and we're trying to fix the purified prep for later analysis. thanks in advance. Bill Ross William (Bill) Ross Director - FACS Core Facility MR-4, Box G005 University of Virginia School of Medicine Charlottesville, VA 22908 office (804) 982-1586 fax (804) 924-1221 e-mail:wgr8w@virginia.edu
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