A rapid micromethod for evaluating T cell subsets in blood using monoclonal antisera

A simple accurate method is described for enumerating T cell subsets in whole blood. The method, which depends on indirect immunofluorescence using biotin-coupled monoclonal antisera and fluorescein-coupled avidin, and propidium iodide for nuclear counterstaining, was compared with the conventional method based on initial separation of lymphocytes by density flotation and exposure to monoclonal antisera. Accurate identification of mononuclear cells in whole blood by nuclear staining with propidium iodide was established. The whole blood method gave numbers for T cell subpopulations generally comparable with those obtained by the conventional method, but slightly higher numbers of Leu2a+ cells were found by the whole blood method, and shown to be higher because of selective loss of Leu2a+ plastic-adherent cells in the conventional method. The whole blood method is quicker, uses only 0.5 ml blood and is economical in use of monoclonal reagents.