Development of nucleic acid extraction and real-time recombinase polymerase amplification (RPA) assay integrated microfluidic biosensor for multiplex detection of foodborne bacteria

Rapid and accurate screening of foodborne bacteria is of great significance for food safety, particularly in the areas where health resources for grass-root service are inadequate. In this study, we fabricated a fully integrated micro-platform (named FID-MP) performing nucleic acid extraction, recombinase polymerase amplification (RPA), and signal detection for point-of-care testing (POCT) of multiplex pathogens. After the on-chip nucleic acid extraction, which adopted a filtration membrane, to obtain the DNA template of bacteria, the purified DNA was driven to the detection chip to conduct nucleic acid amplification. The fluorescent images can be read by using a smartphone under the portable signal detector. Using FID-MP, eight foodborne bacteria were detected with 100% specificity and similar sensitivities as compared to standard real-time RPA reactions. The entire process was accomplished within 60 min. Overall, FID-MP eliminates the need for bulk equipment and skilled personnel, holding promise to become a versatile sample-to-answer platform for foodborne bacteria monitoring.