Lymphocyte subset determination using a hematology analyzer

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Abstract
Anti‐CD4 antibody (T4)‐coated microspheres were used to label CD4 cells in whole blood. The mixture was lysed and analyzed by a modified Coulter VCS hematology analyzer, which differentiated microspherelabeled cells by a change in Coulter volume, conductance, and light scatter. %CD3+/CD4+ fluorescent values from a profile were compared to %CD4 values using the VCS‐microsphere method. CD3 gating was used to exclude CD4+ monocytes from the 9OLS‐FALS lymphocyte gate. The results correlated well (R = 0.996). The percentage of CD4+ lymphocytes from profile scatterplots and VCS scatterplots showed a line of regression close to the equivalence line (n = 76, slope = 0.96) when CD3 gating was used for the profile. These results suggest that CD3 gating, though necessary for 9OLS‐FALS scatterplots, may not be necessary for volume‐conductance‐light scatterplots.