ELISA is commonly used for:
Enzyme-linked immunosorbent assay revolutionized immunodiagnostics by converting antigen antibody binding into measurable enzymatic colorimetric signal without radioactivity, originally described by Eva Engvall and Peter Perlmann 1971. Polystyrene microtiter plate with high binding capacity via hydrophobic interaction adsorbs capture antibody coated overnight at alkaline pH 9.6 carbonate buffer 1 to 10 microgram per ml. Non-specific sites blocked with protein bovine serum albumin five percent, casein, or nonfat milk to prevent background adsorption through hydrophobic pockets. Sample containing antigen such as viral capsid HIV p24, cytokine IL-6, or hormone insulin incubated allowing specific capture, unbound material removed by repeated washes with phosphate buffered saline containing 0.05 percent Tween-20 disrupting weak interactions. Detection uses secondary antibody conjugated covalently to horseradish peroxidase or alkaline phosphatase via periodate oxidation forming Schiff base. Addition of chromogenic substrate tetramethylbenzidine TMB with hydrogen peroxide yields oxidation to blue diimine measured after sulfuric acid stopping as yellow product absorbance 450 nm proportional to analyte concentration via four-parameter logistic standard curve with dynamic range three logs.
Ref: Engvall & Perlman Immunochemistry 1971 ELISA; Alberts MBoC ELISA HRP TMB sandwich principle diagnostic.