Probing antigen-antibody interactions on electrode supports by the biocatalyzed precipitation of an insoluble product

The amplified sensing of an antibody by an antigen monolayer-functionalized transducer by using a biocatalyzed precipitation of an insoluble product o n the transducer is addressed. Faradaic impedance spectroscopy, cyclic volt ammetry and microgravimetric, quartz crystal microbalance analyses are used to probe the precipitation of the insoluble product on the transducer. A d initrophenyl, DNP, antigen monolayer is assembled on a Au-electrode, or a A u-quartz crystal, as a sensing interface for the dinitrophenyl-antibody, DN P-Ab. An anti-Fc-antibody-HRP conjugate is used as a biocatalytic probe for the formation of the antigen/DNP-Ab complex on the transducer. Biocatalyze d oxidation of 4-chloronaphthol by H2O2 in the presence of the anti-Fc-anti body-HRP conjugate yields an insoluble product on the transducer. Formation of the insoluble film on the electrode results in the increase of interfac ial electron-transfer resistances detected by impedance spectroscopy or cyc lic voltammetry. The precipitate formation also results in the mass increas e of the modified Au-quartz crystal detected as a frequency change of the p iezoelectric transducer. The DNP-Ab is easily sensed at a sensitivity that corresponds to 0.5 ng mL(-1) (3 x 10(-12) M).