REAGENTS USED IN MICROPARTICLE-ENHANCED NEPHELOMETRIC IMMUNOASSAY

Microparticle-enhanced nephelometric immunoassay allows antigen quanti tation by measurement of the light scattered by microparticle-antigen conjugates in a competitive inhibition assay. As previously reported, such an immunonephelometric method is based on the agglutination of hy drophilic microparticle-antigen conjugates occurring in the presence o f specific polyclonal antibodies. Using human chorionic gonadotropin a s model, this work studies the main factors affecting microparticle-an tigen conjugate agglutination and its nephelometric quantification. It allows the understanding of some general problems concerning micropar ticle agglutination. The immunonephelometric properties of micropartic le-antigen conjugates varied with the amount of antigen bound to the m icroparticle (in relation to the conditions of the microparticle coati ng) and with the immunoreactivity of the bound antigen molecules (in r elation to the agglutinating reagent used). Free monoclonal antibodies had limited capacities as agglutinating reagent in microparticle-enha nced nephelometric immunoassay based on a competitive inhibition. The mixture of complementary monoclonal antibodies was generally advantage ous for conjugate agglutination and the cooperative effect of antibodi es was maximum in polyclonal antiserum. The use of xenogenic antibodie s did not improve the sensitivity of the inhibition of agglutination, in spite of the lower concentration of specific antibodies necessary t o agglutinate the microparticle-antigen conjugate.