Feby Wijaya Pratiwi, Patsamon Rijiravanich, Mithran Somasundrum, Werasak Surareungchai
We describe a sensitive electrochemical immunoassay for Salmonella enterica serovar Typhimurium, a common foodborne pathogen which can cause infection at extremely small doses. The assay is based on the recognition of DNA biobarcode labels by differential pulse anodic stripping voltammetry (DPASV), following Ag enhancement. The biobarcodes consist of latex spheres (mean diameter 506 nm ± 22 nm) modified by ferromagnetic Fe3O4 particles. Each biobarcode is loaded by adsorption with approx. 27 molecules of mouse monoclonal antibody against S. Typhimurium and 3.5 × 105 molecules of 12 mer ssDNA. The assay is performed by adding the biobarcode, S. Typhimurium cells, and biotin-conjugated rabbit polyclonal antibody against Salmonella into well plates. After antigen-antibody binding, magnetic collection enables the excess polyclonal antibody to be washed off. Exposure to avidin-coated screen printed electrodes, and formation of the avidin-biotin bond, then enables the excess biobarcode to be removed. The biobarcode remaining on the electrode is quantified by DPASV measurement of Ag+ ions following catalytic Ag deposition. The assay showed a negligible response to 107 CFU mL-1E. coli and had a limit of detection of 12 CFU mL-1 in buffer, and 13 to 26 CFU mL-1 for heat-killed and whole cell S. Typhimurium in plain milk, green bean sprouts and raw eggs. To the best of our knowledge, this is the lowest reported limit of detection for Salmonella by an electrochemical immunoassay not requiring sample pre-enrichment. © The Royal Society of Chemistry.
School of Bioresources and Technology, King Mongkut's University of Technology, Thonburi Bang Khun Thian, Bangkok 10150, Thailand; Chemistry Dept, Faculty of Mathematics, Brawijaya University, Malang 65145 East Java, Indonesia; National Center for Genetic Engineering and Biotechnology, National Sciences and Technology Development Agency, King Mongkut's University of Technology, Thonburi Bang Khun Thian, Bangkok 10150, Thailand; Biological Engineering Graduate Program, King Mongkut's University of Technology, Thonburi Bangmod, Bangkok 10140, Thailand
