Strategy for the immobilization of monoclonal antibodies on solid-phase supports.

Using matrices based upon Affi-Gel and Affi-Prep, we have examined conditions during the immobilization of antibodies (immunoglobulin G, IgG) that influence the performance of immunosorbents. Such conditions include: coupling pH, coupling kinetics, antibody density on the immunosorbent and the activation chemistries utilized for the immobilization process. These studies have shown that the capacity for antigen does not increase with increased antibody coupling efficiency. Presumably, increased coupling times or efficiencies lead to multi-site attachment of the antibody to the matrix, thereby causing inactivation. Immunosorbents containing low densities of IgG were found to have greater capacity for antigen on a per mole IgG basis. This suggests steric crowding of antigen at high antibody density. Finally, immunosorbents prepared through IgG carbohydrate linkages (oriented coupling) show dramatic increases in antigen capacity over those prepared by stochastic (random) coupling through IgG primary amino groups. A combination of low IgG density and oriented coupling of the IgG via the carbohydrate moiety may represent the best strategy for the preparation of immunosorbents.