Dear Andy, Human or mouse ? Do they have a "spare" channel for a dump gate ? Can they use a myeloid marker to gate these cells out ? I have used anti-GR-1 in the mouse, which is usually very good as a negative gate for the large, high autoflourescent cells of myeloid/gran lineage. Also, what about a viability stain like PI - those autofl. cells may be stain slightly with PI, even when viable. Good luck, Rachel ======================================================= Rachel M. Gerstein, Ph.D. Department of Molecular Genetics and Microbiology Graduate Program in Immunology/Virology University of Massachusetts Medical School 55 Lake Avenue North Worcester, MA 01655-0002 (508) 856-1044 (508) 856-5920 (FAX) > ---------- > From: Andy Lane > Sent: Tuesday, October 5, 1999 7:48 AM > To: Cytometry Mailing List > Subject: Re: Autofluorescence in bone marrow samples > > > Dear All, > > I'm posting this for a colleague, so don't have all the details, but hope > that someone may have some suggestions. > > They are using "whole blood" bone marrow preparations in analysis of > various > leukaemias/lymphomas, and consistently find a problem with autofluorescent > cells that give signals in both the FL1 and FL2 channels (they have that > typical non-specific diagonal look on a dot-plot of FL1-FL2). The > non-specific signals appear even in the absence of any antibody, so this > seems to be a true autofluorescence rather than non-specific antibody > binding. > > They are reluctant to gate out on FSC/SSC after back-gating as they feel > that they risk losing important cells. Does anyone have a simple way of > overcoming this problem, or even know for certain what the cells are > likely > to be. They are not seen after ficoll separation, and scatter suggests > that > they are of myeloid origin > > > Andy > (andy@serotec.co.uk) > > > > > > >
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