A new quantitative fluorometric binding assay that uses fluoresceinated aggregated IgG is proposed for the study of Fc receptors. The method was compared with a radiolabeling binding assay on three well characterized murine cell lines (38C-13, EL4, and BW). The apparent association constant of the binding and the amount of aggregated IgG bound per cell at saturation were calculated. The fluorometric assay enables the detection of 5 × 10-10 M bound aggregated IgG. Inhibition studies with monomeric IgG, reduced and alkylated aggregated IgG, and aggregated F(ab′)2 fragments of IgG confirmed the specificity of the assay. Staphylococcal protein A inhibited the binding of the aggregated IgG to Fc receptors.
This work was supported in part by the Tyssen Foundation, FKFO, FGWO, ASLK Cancer Fund, Solvay Tournay Fund for Medical Research.