Simultaneous Determination of Absolute Total Lymphocyte and CD4+ Lymphocyte Levels in Peripheral Blood by Flow Cytometry

A distinguishing feature of the Spectrum III flow cytometer is its capacity to analyze a constant volume of cell suspension. The authors have capitalized on this feature to directly and simultaneously measure the absolute numbers of all lymphocytes and CD4+ lymphocytes per microliter of peripheral blood. The study group consisted of 42 hospital patients, 12 former blood donors seropositive for human immunodeficiency virus (HIV), and 12 HIV-seronegative donors. Regression analysis revealed a highly significant correlation (r = 0.97) between Spectrum lymphocyte counts and lymphocyte counts determined by a Coulter Counter S-Plus IV. Similarly, Spectrum CD4 cell counts were significantly correlated (r = 0.98) with CD4 cell counts calculated from the Coulter lymphocyte counts and % CD4+ lymphocytes determined by flow cytometry. These findings indicate that the absolute numbers of lymphocytes and subsets of lymphocytes in peripheral blood can be rapidly and simultaneously measured by flow cytometry. Such an assay should prove useful in studies of HIV infection, where total lymphocyte and CD4 cell levels are important parameters for clinical staging and assessing responses to treatment.