A method for the measurement of the antigen-binding activity of immunoglobulins precipitated in commercially available radial immunodiffusion plates.

A method for determining the antigen-binding activities of specific immunoglobulins in serum is described. Samples of sera were applied to commercial immunodiffusion plates and unprecipitated proteins eluted. The precipitate circles were challenged with specific radio-labelled antigen and exposed to X-ray film. Blackened circles on the radioautograph were found to be good indices of the antigen-binding activity of each immunoglobulin and could be monitored using a Beckman Model R Analytrol. The height of each analytrol peak provided an objective estimate of the intensity of darkening. Variation in the amount of added radio-antigen had a direct effect upon the intensity of the darkened circle at low concentrations. At a given concentration of antigen, however, all antigen-binding sites became saturated and additional increases in antigen concentration did not further increase the density of blackening. The quantity of antigen required to just saturate the combining sites of a particular immunoglobulin is a measure of the antigen-binding activity of that immunoglobulin. The employment and some of the applications of this relatively simple radio-isotopic technique are discussed.