Competitive enzyme immunosensor developed on a renewable carbon paste electrode support

Abstract

A sequential competitive voltammetric immunoassay format performed on a carbon paste electrode surface for the determination of mouse immunoglobulin G (IgG) is described in this work. This device relies on the attachment of goat anti-mouse IgG by adsorption on the electrode surface. An electrochemical pre-treatment is performed with the aim of improving the sensitivity and reproducibility of such immobilisation. The assay comprises first an incubation in the analyte solution and a further reaction with a tracer for the monitoring of the analyte–antibody binding. Mouse IgG labelled with alkaline phosphatase is chosen together with 3-indoxyl phosphate as the enzymatic substrate. Its corresponding product is indigo. This molecule, generated very close to the electrode surface, is insoluble in aqueous solutions and adsorbs on the support. The electrodic oxidation is a reversible process taking place at around −0.4 V (versus Ag/AgCl) in Tris buffer pH 7.2. Alternating current (AC) voltammetry is chosen for its detection. The feasibility of reusing the same carbon paste electrode for several consecutive assays is demonstrated by performing the same electrochemical treatment as done initially. Thus, an effective removal of the protein layer as well as the indigo adsorbed is achieved, obtaining a regenerated surface with adsorptive properties similar to those of freshly prepared electrodes.