KINETICS AND PROPOSED MECHANISM OF THE REACTION OF AN IMMUNOINHIBITION, PARTICLE-ENHANCED IMMUNOASSAY

We report kinetic studies on the reaction of a latex agglutination imm unoassay used to quantify phenytoin in serum. In this assay, polystyre ne particles with a covalently attached analog of phenytoin react with an antiphenytoin monoclonal antibody to form light-scattering aggrega tes, with the rate of this reaction being decreased by addition of phe nytoin from sample. In the absence of free (sample) phenytoin, this re action did not exhibit a maximum rate of agglutination in the presence df excess antibody, i.e., an equivalence point. Furthermore, agglutin ation was inhibitable by free phenytoin even when the latter was added after agglutination of particles with antibody had begun. Most signif icantly, the immunoagglutination proceeded in an identical fashion wit h monovalent F(ab) fragment. These data are consistent with low-affini ty immunospecific particle-antibody complexation, which then induces c olloidal aggregation, without requiring immunospecific bridging by ant ibody molecules. The described mechanism is not generalizable to all l atex agglutination immunoassays, although disturbance of colloidal sta bility may be a component in most assays.