Development of redox-labeled electrochemical immunoassay for polycyclic aromatic hydrocarbons with controlled surface modification and catalytic voltammetric detection.

A redox-labeled direct competitive electrochemical immunoassay for polycyclic aromatic hydrocarbons (PAHs) was developed. A ruthenium tris(bipyridine)-pyrenebutyric acid conjugate was synthesized as the redox-labeled tracer. Its recognition by an anti-PAH monoclonal antibody was confirmed by surface plasmon resonance. In the immunoassay, the antibody was immobilized on (3-glycidoxypropyl)-trimethoxysilane (GPTMS)-modified indium tin oxide (ITO) electrodes. The assay was quantified by measuring the electro-catalytic current of the redox label in an oxalate-containing electrolyte which served as a sacrificial electron donor to amplify the current signal. Formation of GPTMS film on ITO and subsequent antibody immobilization were characterized by X-ray photoelectron spectroscopy (XPS) and electrochemistry. Using a ruthenium tris(bipyridine)-conjugated IgG (IgG-Ru) as the surface-bound redox probe, the highest electrochemical signal was obtained on GPTMS electrodes with 1 h modification. Under the optimized conditions for ITO modification, antibody immobilization and tracer concentration, competition curves for benzo[a]pyrene and pyrenebutyric acid were obtained with a detection limit of 2.4 and 10 ng mL(-1), respectively. The redox-labeled electrochemical immunoassay with signal amplification mechanism offers a potential analytical method for the simultaneous detection of multiple environmental organic pollutants on antibody biochips.