Electrochemical immunosensor signaling by employing enzyme-tagged antibody for the determination of antigen or antibody under single competition reaction format

Target biomolecules in immunosensing, either antigens or antibodies, were detected using competitive reaction with enzyme-tagged antibody. The enzyme-tagged antibody was prepared through covalent conjugation reaction between antibody and glucose oxidase (GOX), as an electrochemical signaling molecule. By adopting dinitrophenyl (DNP) group as a target, the conjugation reaction of GOX-tagged anti-DNP antibody was performed and confirmed by liquid chromatography. The detectable concentration range of GOX-tagged anti-DNP antibody (from 0.1 μg/ml to 0.1 mg/ml) was registered by using electrochemical signaling under competitive reaction. Target biomolecules, including DNP antigen and anti-DNP antibody, were detected by using competitive immuno-reaction with the GOX-tagged anti-DNP antibody on the antigen (DNP)-functionalized sensing surface. Sensing interface was prepared by using dendrimer-assisted self-assembled monolayer. From immunosensor measurements under competition reaction format, target antigen and antibody exhibited linear detection ranges of 200 nM–2 mM and 1 μg/ml–0.1 mg/ml, respectively.