Dear Mark, We have used the J33 clone extensively in a variety of conjugates (FITC, PE and PE:Cy5). It is an extremely reliable reagent and it is a 'recommended reagent' for use in the ISHAGE CD34+ cell enumeration Guidelines. Have you compared this reagent with any other pan-CD45 antibodies, such as HLE-1 (BD) or KC56 (Coulter) to see if you detect similar events with these clones? Have you sorted these CD45- 'events' and had a look at the morphology of them on the microscope? How numerous are they and are they always present in different samples? We do not usually see significant numbers of J33- events in our analyses. Perhaps some of them are nucleated RBCs or megakaryocyte fragments. Such particles and small platelet aggregates can exhibit the light scatter characteristics of the events that you describe. Do you have this problem in fresh samples only, or do you only find them in apheresis concentrates that have been 'sitting around' for a while? Hope this is helpful, Rob Sutherland ISHAGE Stem Cell Enumeration Committee, Toronto Hospital
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