Nanogold-enhanced graphene nanosheets as multienzyme assembly for sensitive detection of low-abundanceproteins

Abstract

A new electrochemical immunosensing protocol was designed for detection of carcinoembryonic antigen (CEA, as a model protein) by using graphene-carried poly(o-phenylenediamine)/gold hybrid nanosheets (GNPGs) as signal tags on the hierarchical dendritic gold microstructures (HDGMs)-modified glassy carbon electrode. To prepare the signal tags, poly(o-phenylenediamine) molecules were initially immobilized on the surface of graphene nanosheets via the π-stacking interaction. Then gold nanoparticles were assembled onto the poly(o-phenylenediamine)-modified graphene nanosheets, which were used for the labeling of anti-CEA detection antibodies and horseradish peroxidase (HRP). The as-prepared GNPGs were characterized by using atomic force microscopy (AFM), transmission electron microscopy (TEM) and UV–vis absorption spectroscopy. The assay was carried out with a sandwich-type immunoassay format in pH 5.5acetic acid-buffered saline solutions containing 2.5mmolL−1 H2O2. Under optimal conditions, the electrochemical immunoassay exhibited a wide dynamic range of 0.005–80ngmL−1 toward CEA standards with a low detection limit of 5.0pgmL−1. Intra- and inter-assay coefficients of variation were less than 11.5%. No significant difference at the 0.05 significance level was encountered in the analysis of 6clinical serum specimens and 6spiked blank new born cattle serum specimens between the developed immunoassay and commercially available electrochemiluminescent (ECL) method for the detection ofCEA.