Development of a High Sensitivity Rapid Sandwich ELISA Procedure and Its Comparison with the Conventional Approach

A highly sensitive and rapid sandwich enzyme-linked immunosorbent assay (ELISA) procedure was developed for the detection of human fetuin A/AHSG (alpha 2-HSglycoprotein), a specific biomarker for hepatocellular carcinoma and atherosclerosis. Anti-human fetuin A antibody was immobilized on aminopropyltriethoxysilanemediated amine-functionalized microtiter plates using 1-ethyl-3[3-dimethylaminopropylicarbodiimide hydrochloride and N-hydroxysulfosuccinimide-based heterobifunctional cross-linking. The analytical sensitivity of the developed assay was 39 pg/mL, compared to 625 pg/ mL for the conventional assay. The generic nature of the developed procedure was demonstrated by performing human fetuin A assays on different polymeric matrixes, i.e., polystyrene, poly(methyl methacrylate), and polycyclo-olefin (Zeonex), in a modified microliter plate format. Thus, the newly developed procedure has considerable advantages over the existing method.