Electrochemical Immunosensor for the Detection of Staphylococcal Enterotoxin B (SEB) using Screen-Printed Electrodes (SPEs)

Abstract

Staphylococcal enterotoxin B (SEB) is responsible for large number of food poisoning cases throughout the world. SEB is an exotoxin and it is one of the several harmful substances produced by the bacterium Staphylococcus aureus. Therefore, it is required to develop methods for the sensitive, reliable, reproducible, cheaper, easy to use and rapid detection of SEB. An electrochemical immunosensor was studied for the fast detection of SEB using disposable screen-printed electrodes. Ascorbic acid-2-phosphate (AA-2P) was used as a new substrate for the voltammetric detection of SEB. In the alkaline buffer solution the alkaline phosphate (ALP) enzymatic hydrolysis product of AA-2P is ascorbic acid (AA). Ascorbic acid is an electroactive substance and gives differential pulse voltammetric (DPV) oxidative response at +380mV (versus Ag/AgCl). The potential +0.39V corresponded to the oxidation of AA. Indirect sandwiched enzyme linked immunosorbent assay (ELISA) is used for the detection of SEB. In this method, anti-rabbit IgG for SEB (capturing antibody) was first immobilized on the surface of SPE. These antibodies were further reacting with SEB (antigen) to form antigen-antibody complex. After that, anti-mice IgG for SEB (secondary antibody) were added, followed by ALP-conjugated anti-mice IgG (revealing antibody). The optimal conditions for ALP enzymatic reaction and the volumetric detection were optimized. It was found that the response of voltammetric immunosensor is proportional to the SEB concentration in the range 0.1-100ng/mL. The detection limit was found to be 0.1ng/mL.