Flow-Injection Enzyme Immunoassay for Human IgG by Using Enhanced Chemiluminescence Reaction for Horseradish Peroxidase Label Quantitation

Abstract

Abstract A flow-injection sandwich enzyme immunoassay for human IgG as model antigen by using horseradish peroxidase as label, polystyrene beads as solid support, and the enhanced chemiluminescence reaction for peroxidase quantitation is described. the kinetics of antigen—immobilized antibody interaction has been studied and the quantitative time-concentration ranges of reactions have been estimated. Each of the two immunochemical steps of analysis have been pursued in the kinetic regime. the time for each immunochemical step was reduced to 2–3 min. the enhanced luminescent reaction involving luminol and p-iodophenol as substrates was used to detect the peroxidase label. the conditions for chemiluminescent reaction were optimized. the detection limit for peroxidase in a 3 min assay was 5–10−16 moles/tube. the detection limit for IgG, in the developed immunoassay, is 10−9 M, the overall time of the assay being 5–10 min.