Quantitation of mouse immunoglobulin allotypes by a modified solid-phase radioimmune assay

Allotype-reactive polystyrene tubes were made by first coating their inner surface with immunglobulin (Ig) solutions containing the allotype of interest; this was followed by the application of anti-allotype antibody. The latter was covalently cross-linked to the adsorbed allotype antigen by means of glutaraldehyde to prevent the dissociation of antibody from the surface of the tube in subsequent steps (glutaraldehyde treatment did not affect the specificity of these tubes). Under conditions of antibody excess, allotype-reactive tubes were treated next with 125I-labeled 7S Ig from standard solutions in order to estimate the concentration of Ig allotypes in these solutions. Then, to determine serum allotype concentrations, we used the standard solutions and the serum unknown to carry out competition experiments between labeled and unlabeled allotype antigen. Quantitation of serum Ig allotypes in this way featured a high signal to noise ratio (30 to 100), an error estimate of 3 to 7%, and sensitivity in the nanogram range.