Gold-film array-electrode for electrochemical ELISA

A gold-film array-electrode (8-channel) has been developed for application to an electrochemical ELISA system. The principle of the electrochemical ELISA is the amperometric quantification of oxidized 3,5,3′,5′-tetramethylbenzidine (TMB) which is widely used as a chromogenic substrate of horseradish peroxidase (HRP) in commercial ELISA kits. To make an 8-channel array-electrode, eight sets of three-electrode systems were integrated with the spacing of a 96-well microplate. The gold-film electrode for working- and auxiliary-electrode was made by sputtering a gold layer on a polyetherimide (PEI) strip, and the reference electrode was made by spin coating with Ag/AgCl paste. Using the gold-film electrode, the measurement parameters, such as analysis time, electrode area, measuring temperature and so on, were optimized. Using 50 array-electrodes, the electrode-to-electrode difference in sensitivity was estimated to be 8.2%. To demonstrate the practical applicability of the gold-film electrode, it was applied to commercial ELISA kits for the detection of HIV (antibody), HBV (antigen and antibody), and HCV (antibody). The life time of the gold-film electrode was estimated for use for over 500 repetitive measurements.