Ionic liquids-doped organic-inorganic hybrid film for electrochemical immunoassay for hepatitis B surface antigen

A new mediator-free amperometric immunoassay for hepatitis B surface antigen (HBsAg) in human serum was designed by means of immobilizing horseradish peroxidase-hepatitis B surface antibodies conjugates (HRP-HBsAb) on ionic liquids-doped organic-inorganic hybrid film. The composite film including magnetic nanogold particles and nanoalumina particles provides a friendly microenvironment for the immobilization of biomolecules. The presence of ionic liquids enhances the electron communication between the immobilized biomolecules and the base electrode and improves the sensitivity of the electrochemical immunoassay. With a non-competitive immunoassay format, the formation of the immunocomplex between the immobilized HRP-HBsAb and HBsAg in sample solution exhibited a barrier of direct electrical communication between the immobilized HRP and the base electrode and changed the bioelectrocatalytic properties of the immobilized HRP towards H₂O₂ in the detection solution. Under optimal conditions, the developed immunosensors displayed a good current response in a dynamic range from 1.2 to 430 ng/mL with a limit of detection of 0.3 ng/mL HBsAg (at 3s). The proposed immunosensors have good precision, high sensitivity, acceptable stability, and reproducibility and could be used for the HBsAg detection in human serum with consistent results in comparison with those obtained by the enzyme-linked immunosorbent assay method.