EVALUATION OF A FLOW CYTOMETRIC METHOD FOR SIMULTANEOUS LEUKOCYTE PHENOTYPING AND QUANTIFICATION BY FLUORESCENT MICROSPHERES

We describe a flow cytometric method using a newly designed product, f luorochrome-containing microspheres (Flow Count fluorospheres), which facilitates the precise quantification of cells in whole blood samples or heterogeneous cell suspensions on a single-cell level. These micro particles are easily distinguishable from other events of interest and can be detected by their light-scattering and fluorescence properties . In contrast to the traditional manual or automated cell-counting tec hniques, this method offers the opportunity to quantify cells simultan eously with flow cytometric immunophenotyping without additional cell loss or other cell preparation steps, We evaluated the accuracy and re producibility of this flow cytometric method on the determination of C D45+ leukocyte counts and compared the results with those obtained by conventional techniques. Particular interest was focused on the behavi or of cells and fluorospheres regarding their sedimentation rate over the period of analysis. The data from 48 blood samples with low, norma l, and high leukocyte counts confirmed the reliability and comparabili ty of the flow cytometric method, permitting the determination of whit e blood cell concentration at least to a Limit of 100 cells/mu l. A br oad field of applications will benefit from this flow cytometric suppl ement because it is easy to perform and highly accurate. The results a ppear to be transferable to clinical decision-making monitoring of CD4 + lymphocytes Ln patients infected with human immunodeficiency virus o r of CD34+ hematopoietic cells, optimizing the harvest for peripheral blood stem cell transplantation. Cytometry 33:310-317, 1998, (C) 1998 Wiley-Liss, Inc.