Laurie, I have some experience with platelet analysis on BD FACScan. BD has a protocol, technical bulletin #Flow Cytometric Analysis of Platelets. Briefly, use log-FCS and log-SSC for acquisition. This places platelets in center of plot in a diagonal cluster, with WBC and RBC in upper right corner. We did not fix our samples, stained and acquired fresh. Also we use 1% goat serum in PBS as diluent/blocking agent, as we find less autofluorescence with gost than FCS or albumin. If you are using whole blood, I would suggest collecting in ACD-A ~1:7, as this lowers the pH to 6.8-6.9 and prevents any aggregation. Also if collecting from anticubical, use large bore needle, with gentle collection (DO NOT use vacutaineer), and discard first 2ml as this will contain activated platelets. Hopefully this will be of some help. April Translational Research & Flow Cytometry Blood & Marrow Transplantation Univ TX MD Anderson Cancer Center
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