Mouse monoclonal anti rabbit IgG coupled to monodisperse polymer particles. Comparison with polyclonal antibodies in radioimmunoassay for thyroid hormones

Abstract

A solid phase second antibody was prepared by covalent coupling of a mouse monoclonal anti rabbit IgG to monodisperse particles. This preparation was compared with immunosorbent purified sheep anti rabbit IgG antibodies coupled to the same particles. The monoclonal antibody bound rabbit IgG with a dissociation constant of 3 X 10(-11) L/mol, and the binding was Fc specific. The sheep antibodies had a similar Kd and about 75% of the activity was directed against the Fc portion of IgG. The binding capacity per mol of both solid phase antibodies was 0.7 mol of rabbit IgG. Monoclonal and polyclonal solid phase antibodies were equally effective as separating agents in various radioimmunoassays. Direct coupling of the rabbit antibodies to the solid phase resulted in a marked loss of binding capacity for the respective thyroid hormones. However, when rabbit anti-thyroxine or anti-triiodothyronine were preadsorbent to second-antibody-coated particles the binding capacities of the former antibodies were well preserved.