Simultaneous electrochemical immunosensing of alpha-fetoprotein and prostate specific antigen using a glassy carbon electrode modified with gold nanoparticle-coated silica nanospheres and decorated with Azure A or ferrocenecarboxylic acid

We describe an electrochemical immunosensor for the simultaneous determination of alpha-fetoprotein (AFP) and prostate specific antigen (PSA) via a modified glassy carbon electrode. Silica nanoparticles (200–300 nm i.d.) with good monodispersity and uniform shape were synthesized, and the following species were then consecutively immobilized on their surface: gold nanoparticles (AuNPs; 5–15 nm i.d.), secondary antibody (Ab₂) and the redox-probes Azure A or ferrocenecarboxy acid (Fc). In parallel, two types of primary antibodies (Ab₁) were co-immobilized on the surface of the dissolved reduced graphene oxide sheets (rGO) that were also decorated with AuNPs. In the presence of antigens (AFP or PSA), the Ab₂/Si@AuNPs carrying Azure A and Fc are attached to the AuNP/rGO conjugate via a sandwich type immunoreaction. Differential pulse voltammetry (DPV) was employed to measure the resulting changes in the signal of Fc or Azure A. Two well-resolved oxidation peaks, one at ?0.48 V (corresponding to Azure A) and other at?+?0.12 V (corresponding to Fc; both vs. SCE) can be observed in the DPV curves. Under optimal conditions, AFP and PSA can be simultaneously determined in the range from 0.01 to 25 ng mL￣¹ for AFP, and from 0.012 to 25 ng mL￣¹ for PSA. The detection limits are 3.3 pg mL￣¹ for AFP and 4.0 pg mL￣¹ for PSA (at a signal-to-noise ratio of 3). The method was applied to (spiked) real sample analysis, and the recoveries are within 96.0 and 107.2 % for PSA, and within 100.9 and 105.8 % for AFP, indicating that this dual immunosensor matches the requirements of clinical analysis.

(A) Two types of signal labels preparation process. (B) The immunosensor preparation and detection process.