Graphene-based immunosensor for electrochemical quantification of phosphorylated p53 (S15).

We reported a graphene-based immunosensor for electrochemical quantification of phosphorylated p53 on serine 15 (phospho-p53(15)), a potential biomarker of gamma-radiation exposure. The principle is based on sandwich immunoassay and the resulting immunocomplex is formed among phospho-p53 capture antibody, phospho-p53(15) antigen, biotinylated phospho-p53(15) detection antibody and horseradish peroxidase (HRP)-labeled streptavidin. The introduced HRP results in an electrocatalytic response to reduction of hydrogen peroxide in the presence of thionine. Graphene served as sensor platform not only promotes electron transfer, but also increases the surface area to introduce a large amount of capture antibody, thus increasing the detection sensitivity. The experimental conditions including blocking agent, immunoreaction time and substrate concentration have been optimized. Under the optimum conditions, the increase of response current is proportional to the phospho-p53(15) concentration in the range of 0.2-10 ng mL(-1), with the detection limit of 0.1 ng mL(-1). The developed immunosensor exhibits acceptable stability and reproducibility and the assay results for phospho-p53(15) are in good correlation with the known values. This easily fabricated immunosensor provides a new promising tool for analysis of phospho-p53(15) and other phosphorylated proteins.