Characterization of antigen-binding receptors in vitro. I. An equilibrium method for measuring antigen binding in murine cells

Abstract

This study reports a novel method for studying the binding of soluble antigen (bovine serum albumin, BSA) to surface receptors on lymphoid cell populations under equilibrium conditions in the presence of 10% normal rabbit serum or 20 mg/ml of ovalbumin. Virtually no nonspecific uptake was demonstrated to nonlymphoid tissues. Detectable quantities of BSA could be found on both nonimmune and immune lymphoid populations. The binding of BSA was demonstrated to be antigen specific and to be proportional to the numbers of binding cells. The quantity of antigen bound was proportional to the free antigen exposed to the population and saturation could be achieved with 1 to 2 microgram of antigen/1.2 ml of culture, per 10 X 10(7) cells. The kinetics of antigen binding was very rapid and occurred within 10 min at 4, 27, and 37 degrees C. The binding was independent of the viability of cells. Binding was antigen specific and could be partially blocked by cross-reacting serum albumins, but not by non-cross-reacting albumins. Binding was independent of cytophylic antibody concentrations in the serum.