Sensitive electrochemical immunoassay of carcinoembryonic antigen with signal dual-amplification using glucose oxidase and an artificial catalase

A new dual-amplification strategy of electrochemical signal based on the catalytic recycling of the product was developed for the antigen–antibody interaction by glucose oxidase-conjugated gold–silver hollow microspheres coupled with an artificial catalase, Prussian blue nanoparticles, on a graphene-based immunosensing platform. [Display omitted] ► We designed an electrochemical immunoassay of carcinoembryonic antigen with signal dual-amplification. ► A new signal tag with glucose oxidase-conjugated gold–silver hollow microspheres. ► An artificial catalase, Prussian blue nanoparticles, on a graphene-based sensing platform. ► Comparative study of the electrochemical immunoassay by using various signal tags and sensing platforms. A new dual-amplification strategy of electrochemical signal based on the catalytic recycling of the product was developed for the antigen–antibody interaction by glucose oxidase (GOD)- conjugated gold–silver hollow microspheres (AuAgHSs) coupled with an artificial catalase, Prussian blue nanoparticles (PB), on a graphene-based immunosensing platform. The first signal amplification introduced in this study was based on the labeled GOD on the AuAgHSs toward the catalytic oxidation of glucose. The generated H 2O 2 was catalytically reduced by the immobilized PB on the graphene nanosheets with the second amplification. With a sandwich-type immunoassay format, carcinoembryonic antigen (CEA) was monitored as a model analyte by using the synthesized AuAgHSs as labels in pH 6.0 phosphate buffer containing 10 mM glucose. Under optimal conditions, the electrochemical immunosensor exhibited a wide dynamic range of 0.005–50 ng mL −1 with a low detection limit (LOD) of 1.0 pg mL −1 CEA (at 3 σ). Both the intra- and inter-assay coefficients of variation (CVs) were lower than 10%. The specificity and stability of the immunosensor were acceptable. In addition, the assay was evaluated for clinical serum specimens, and received a good correlation with those obtained by the referenced electrochemiluminescent (ECL).