Phenotypic characterization of individual interferon-gamma-producing cells after OKT3 antibody activation.

This is the first study using a direct approach to determine the phenotype of interferon gamma (IFN-gamma)-producing cells after polyclonal T cell activation. Blood mononuclear cells (MNC) were cultured and stimulated by OKT3 antibody to produce IFN-gamma. They were then stained with a panel of monoclonal antibodies with specificity for cell surface antigens, fixed and permeabilized in suspension and stained for cytoplasmic IFN-gamma using monoclonal antibodies and indirect immunofluorescence technique. IFN-gamma appeared mainly as a local, polar accumulation in the cytoplasm in a juxtanuclear position, probably identical to the Golgi apparatus. After a culture period of 20 h, 6-11% and 0.2-0.6% of the OKT3-activated MNC from adults or newborns, respectively, produced IFN-gamma in cultures. Unstimulated cultures contained maximally 0.1% IFN-gamma-positive MNC. The majority of the IFN-gamma-producing MNC were T cells and expressed the T11 antigen and receptors for interleukin 2. To our surprise most of these T cells also carried receptors for complement 3bi (OKM1) but did not express class II molecules. Less than 50% displayed T4 or T8 antigens in three out of the four experiments performed. Natural killer cells, as recognized by Leu 7 or B73.1 antibodies, contributed only marginally to the IFN-gamma synthesis whereas B cells (OKB7) and monocytes (Leu M3) showed no production of IFN-gamma.