Flow cytometric studies of the binding of monoclonal antibodies OKT3, OKT4 and OKT8.

The binding of monoclonal antibodies (OKT3, OKT4 and OKT8) to human T cells was investigated by flow cytometry. A flow cytometer was calibrated with standard fluorescence microspheres, which permitted quantitation of the number of bound antibody molecules. Considerable care was taken to perform the flow cytometric assay at a constant temperature and the effect of temperature on the binding reaction was examined. The binding of OKT3, OKT4 and OKT8 exhibited saturation kinetics. The maximum binding varied with temperature. Kinetic analysis according to the Hill equation revealed that the value of the Hill coefficient for OKT3 changed from 1.8 to 1.0 when the temperature was raised from 12 degrees C to 36 degrees C, whereas the corresponding values for OKT4 and OKT8 did not vary with temperature. Thermodynamic functions obtained from the Van't Hoff plot showed that the binding of OKT3 was exothermic whereas the binding of OKT4 and OKT8 were endothermic.