Influence of specificity and affinity of antibodies on the kinetics of a sandwich enzyme immunoassay for human IgG.
In comparative experiments, we desired to determine the influence of specificity and affinity of various combinations of antibodies on the kinetics of a "sandwich" enzyme immunoassay for human IgG. Some antibodies were immobilized and others labeled with glucose oxidase. When antibodies employed were heterogeneous in specificity and affinity, long periods of time were required to reach equilibrium, especially on the second step of the assay. In contrast, when antibodies were separated into populations with specificity towards two different antigenic sites, and each one was used as immunoadsorbent and labeled, time was significantly reduced. Finally, when the assay took place--the last antibodies being selected by their high affinities--the rate of the reaction improved even more. We assume selection of antibodies by specificity and affinity by two antigenic sites as an essential requirement to improve performance of these assays, independently of the antigen and marker used.
D000818:Animals; D000888:Antibodies, Anti-Idiotypic / Q000032:analysis; D000915:Antibody Affinity; D000918:Antibody Specificity; D001666:Binding Sites, Antibody; D003429:Cross Reactions; D006801:Humans; D007124:Immunoenzyme Techniques; D007074:Immunoglobulin G / Q000032:analysis* / Q000276:immunology; D007143:Immunoglobulin Heavy Chains / Q000276:immunology; D007700:Kinetics; D011817:Rabbits
D000888:Antibodies, Anti-Idiotypic; D007074:Immunoglobulin G; D007143:Immunoglobulin Heavy Chains