Synthesis and Characterization of a Novel All-in-One Graphene Oxide-Nafion Polymer Bioconjugate for Application in Electrochemical Biosensing of the <i>Opisthorchis viverrini</i> Antigen

Bioconjugates in electrochemical biosensors can significantly enhance the detection process and sensitivity. In this study, we synthesized a monoclonal antibody-conjugated nanocomposite of graphene oxide and Nafion (GO-Nf-mAb) for application in an electrochemical biosensor as a novel all-in-one bioreceptor. The incorporation of Nafion (Nf) improved the stability, dispersity, and antibody immobilization on the GO surface, thereby increasing the sensitivity of the biosensor. The impact of Nafion on GO stability and antibody conjugation was thoroughly investigated and compared to that of Nafion-free conjugation via various characterization techniques, including X-ray photoelectron spectroscopy (XPS) and synchrotron radiation near-edge X-ray absorption fine structure. The presence of Nafion during monoclonal antibody (mAb) conjugation resulted in an increased peak intensity of the NH₂ band in XPS analysis and the highest intensity of C=O groups in O-K edge analysis, indicating a greater yield of the antibody. This innovative electrochemical biosensor exhibited a low detection limit of 1.68 ng mL^-1 in spiked urine, a wide linear range, and high reproducibility, outperforming conventional detection methods for <i>Opisthorchis viverrini</i> (OV) antigen detection. Our developed electrochemical biosensor introduces a novel and straightforward fabrication process using an all-in-one bioconjugate that serves as a bioreceptor, transducer, and blocking reagent simultaneously. Overall, this study offers a new insight on Nafion application in bioconjugation, and the GO-Nf-mAb conjugate-based electrochemical biosensor promises high sensitivity and a hassle-free immobilization process for OV antigen quantification.