STUDIES ON AFFINITY CONSTANTS OF HAPTEN-SPECIFIC MONOCLONAL-ANTIBODIES USING ANTIBODY-IMMOBILIZED ELISA

An enzyme-linked immunosorbent assay to measure affinity constants of hapten-specific monoclonal antibodies has been developed. In this meth od an antibody-immobilizing format is used. Antibody affinity toward h apten-enzyme tracer is first calculated by using Scatchard's method. U sing this affinity, antibody-hapten affinity is then calculated by fol lowing the principles of chemical thermodynamics and mass conservation . The affinity from this method agrees with the IC50 values in ELISA. This method is useful in early determination of affinity during monocl onal antibody development. Compared with the antigen-coated format, th is method avoids the problem of distinguishing binding of unliganded a ntibody and singly liganded antibody to antigens on solid phase.