In the present study, a simple and rapid multiplexed bead-based mesofluidic system (BMS) was developed for simultaneous detection of food-borne pathogenic bacteria, including Staphylococcus aureus, Vibrio parahaemolyticus, Listeria monocytogenes, Salmonella, Enterobacter sakazakii, Shigella, Escherichia coli O157:H7, and Campylobacter jejuni. This system is based on utilization of isothiocyanate-modified microbeads that are 250 m in diameter, which were immobilized with specific amino-modified oligonucleotide probes and placed in polydimethylsiloxane microchannels. PCR products from the pathogens studied were pumped into microchannels to hybridize with the oligonucleotide-modified beads, and hybridization signals were detected using a conventional microarray scanner. The short sequences of nucleic acids (21 bases) and PCR products characteristic of bacterial pathogens could be detected at concentrations of 1 pM and 10 nM, respectively. The detection procedure could be performed in less than 30 min with high sensitivity and specificity. The assay was simple and fast, and the limits of quantification were in the range from 500 to 6,000 CFU/ml for the bacterial species studied. The feasibility of identification of food-borne bacteria was investigated with samples contaminated with bacteria, including milk, egg, and meat samples. The results demonstrated that the BMS method can be used for effective detection of multiple pathogens in different foodstuffs.Bacterial food-borne pathogens pose a significant threat to human and animal heath. These organisms are the leading causes of illness and death in less developed countries, killing approximately 1.8 million people annually, and they are the third leading cause of death. In developed countries, foodborne pathogens are responsible for millions of cases of infectious gastrointestinal diseases each year, costing billions of dollars in medical care and lost productivity (42). The Centers for Disease Control and Prevention estimates that food-borne diseases cause approximately 76 million illnesses, 325,000 hospitalizations, and 5,000 deaths in the United States each year (25). The Food and Drug Administration's 2005 Food Code states that the estimated cost of food-borne illness is $10 billion to $83 billion annually (29). The major food-borne pathogens include Salmonella, Listeria monocytogenes, Staphylococcus aureus, Vibrio parahaemolyticus, Campylobacter jejuni, Escherichia coli O157:H7, Enterobacter sakazakii, and Shigella (14,32,41).Rapid detection and identification of pathogens and other microbial contaminants in food are needed by the food industry, food safety agencies, and public health bureaus. Traditional methods to detect food-borne bacteria often rely on time-consuming growth in culture media, isolation of bacteria, biochemical identification, and sometimes serology (5, 8). There have been many attempts to develop faster, more convenient, more sensitive, and more specific techniques for detection and diagnosis of pathogenic bacteria, including immunological m...