Amperometric Detection of Staphylococcal Enterotoxin B in milk using Magnetic Bead-Based Immunoassay on Disposable Screen Printed Carbon Electrode Strip
S.K. Rastogi, S. Gibbon, J.R. Branen, A.L. Branen
University of Idaho, US
Keywords: SEB, milk, SPCEs, Magnetic bead, TMB/Poly80-HRP, ameprometry
Abstract:A novel approach for the sensitive detection of Staphylococcal enterotoxin B (SEB) in milk is reported. The method used sandwich enzyme-linked immunoassay with SEB specific antibodies on magnetic beads. A small cylindrical magnet was fixed, behind the working electrode of the screen-printed carbon electrode (SPCEs) strip. The first SEB-specific antibody was covalently linked with tosylated magnetic beads and used to capture SEB protein from milk samples. The biotin labeled SEB-specific second antibody conjugated with streptavidin labeled poly80 horseradish peroxidase (poly80-HRP) enzyme was served as the signal antibody. The electro-chemical signal were generated using enhanced 3,3’,5,5’-Tetramethylbenzidine (TMB) substrate at a potential +150 mV vs. silver (Ag) reference electrode. The detection limit increases significantly by using poly80-HRP labeled second antibody compared to HRP labeled antibody system. The estimated lowest detection of SEB concentration was found to be 3 pg/ml corresponding to 300 fg in a 100 Bl sample. The results were reproduced several times on different SPCEs and also on same SPCE, which offers future promise for the rapid, specific and cost-effective detection of toxins.