Rapid enumeration of T lymphocytes by a flow-cytometric immunofluorescence method

Combined with current advances in microprocessor-controlled flow cytometers, monoclonal antibodies provide a rapid means of phenotyping individual cell surface markers for a large number of clinical samples accurately and reproducibly, which may provide useful information in diagnosing disease and monitoring patients. We have developed a one-step flow-cytometric immunofluorescence procedure for enumerating E-rosette lymphocytes from whole blood by using the monoclonal antibody OKT11. This antibody recognizes the sheep erythrocyte receptor on the lymphocyte surface and can block sheep E-rosette formation. The flow cytometer we use, an Ortho Spectrum III, distinguishes lymphocytes from other leukocytes by measuring the narrow forward and right-angle light-scattering properties of the cells. The instrument further differentiates T lymphocytes frm non-T lymphocytes by measuring the green fluorescence signal of the OKT11-positive lymphocytes. In a typical sample, 1500--2500 lymphocytes are counted in 25 s. In a study of 158 patient samples, ranging from 1% to greater than 90% E-rosette-positive lymphocytes, the correlation coefficient between the manual E-rosette count and the flow immunofluorescence measurement is 0.943.