Apart from probably non-specific binding via some of the fluorochromes there is always the possibility of cell cell aggregation, especially in non-healthy people. The numbers of cells that aggregated seem to be higher in preparation methods that involve pelleting steps. I had a poster about this on Toronto a few years ago. As these days the numbers of plots displayed is not limiting any more I would always suggest to look at log side scatter versus each of the fluorescence channels used as a basic form of display for general sample assessment. Log side scatter allows to visualise lymphocytes, monocytes neutrophils and eosinophils (and sometimes a few more things) in particular after FACSlyse . Aggregates between monocytes and lymphocytes are easily identified and in most cases there will also be some granulocytes showing for example CD19 as well. In the case I reported it was interesting that the coaggregation was abolished using anti CD45. Thus the classical confirmation of the lymphocyte scatter gate became completely invalid. Regards Gerhard -----Original Message----- From: Annuska Glas [SMTP:aglas@vmresearch.org] Sent: Tuesday, March 07, 2000 11:11 PM To: Cytometry Mailing List Subject: CD14 and CD19 positive cells Hello, I was wondering if anyone has seen a population positive for both CD14 and CD19. I am staining peripheral blood lymphocytes and am looking at the B cell population. I originally just used CD19 to get the B cells. However recently I began using CD3, CD14 to exclude any T cells and monocytes. I will use CD56 as well to exclude NK cells. However, I did see some cells that were positive for both CD19 and CD14. Are these cells monocytes or are they B cells. I do want to look at all B cells, my question is how can I find out if these cells are truly B cells or if they are monocytes? Thanks, Annuska Glas
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