Improving the Sensitivity of Fluorescent Microsphere Immunoassays for the Simultaneous Detection of Foodborne Pathogens Using Dimethylacetamide

The liquid-chip assay is a new method for detecting bacterial surface antigens. This assay conjugated self-prepared monoclonal antibodies against Escherichia coli O157:H7, Listeria monocytogenes, Staphylococcus aureus, and Salmonella enterica serovar Typhi with carboxylated fluorescent microspheres based on the double-antibody sandwich principle. This experiment used dimethylacetamide (DMAC) as the solvent to dissolve 1-ethyl-3-(3-dimethylaminopropyl)-carbodiimide hydrochloride (EDC) and N-hydroxysulfosuccinimide (S-NHS) during conjugation. The modified liquid-chip assay was used to simultaneously detect the four foodborne pathogens. The sensitivity of the assay using the new conjugation method was also evaluated. The limits of detection for E. coli O157:H7, L. monocytogenes, S. aureus, and S. typhi during multiplex detection using the improved method were 0.25, 0.25, 0.5, and 0.25 cfu/mL, respectively, whereas those using the traditional method were 0.5, 0.5, 1, and 0.5 cfu/mL, respectively. Therefore, the improved method is reliable and effectively improves the detection sensitivity of liquid-chip assays.