Solid-phase radioassay for the determination of binding activity of solubilized tritiated Fcγ receptor of guinea pig peritoneal macrophages

Abstract

The reductive methylation procedure for proteins in solution has been adapted for tritium radiolabelling of surface components of macrophages, including the Fcγ receptor. High specific radioactivity of the receptor was obtained with no loss of IgG binding activity, tested by the solid-phase radioassay. To prepare the support for the assay, cellulose filter paper was CNBr-activated and a diamine hydrocarbon reagent was attached as the spacer chain. Next, TNP-hapten residues were covalently coupled to free amine groups. Antibody-coated TNP-paper discs were used as the solid phase for the competitive binding radioassay for the determination of IgG-binding activity of purified Fcγ receptor. The developed radioassay permits the detection of low affinity interactions and/or low quantities of Fc receptors.