Sensitive electrochemical detection of vaccinia virus in a solution containing a high concentration of L-ascorbic acid

Washing processes cannot fully remove interfering species that remain on biosensing surfaces when a sample solution contains a high concentration of interfering species. This study reports an immunosensing scheme employing electroreduction-based electrochemical-chemical (EC) redox cycling that allows sensitive detection of vaccinia virus (VV) in a solution containing a high concentration of L-ascorbic acid (AA). To obtain high signal amplification, an enzymatic reaction by beta-D-galactosidase (Gal) is combined with electroreduction-based EC redox cycling by an oxidant. Among the four possible oxidants (KIO3, NaClO, Ag2O, and H2O2), KIO3 shows the highest signal-to-background ratio and is chosen. During an incubation period of 10 min, Gal converts beta-D-galactopyranoside into p-aminophenol (AP), which is oxidized to p-quinone imine (QI) by KIO3. When -0.05 V vs. Ag/AgCl is applied to an immunosensing electrode, QI is reduced to AP, and the regenerated AP is then reoxidized by KIO3. The electroreduction-based EC redox cycling is induced. An indium-tin oxide electrode modified with reduced graphene oxide and an applied potential of -0.05 V are used to achieve low and reproducible background currents, slow O-2 reduction, and fast electroreduction of QI. KIO3 favorably converts AA into noninterfering species during the incubation period. The detection limit for VV in commercial 50% mandarin juice (AA concentration = 0.7 mM) is 4 x 10(3) plaque-forming unit (PFU) per mL. The new EC redox cycling scheme is promising for sensitive detection of proteins, viruses, and bacteria in solutions containing high concentrations of AA.