On-chip parallel detection of foodborne pathogens using loop-mediated isothermal amplification
Inexpensive, sensitive and specific detection of infectious agents is a critical milestone for preventing disease spread in both the developed and underdeveloped worlds. Rapid and accurate detection of pathogens is especially important for control and treatment of foodborne diseases. According to estimates issued by the Center for Disease Control and Prevention, one out of six Americans will be sick during this year due to consumption of contaminated products and there will be 50,000 related hospitalizations. A point-of-care device capable of performing nucleic acid amplification will enable effective detection of infectious agents in multiple settings, allowing better enforcement of food safety regulations. Current protocols and practices for control of agents responsible for foodborne disease will become cheaper, faster and easier to perform with the incorporation of such devices. Product recalls and food-related diseases will be reduced with a portable system that allows producers, retailers, and consumers to detect pathogens in food samples. In this work, we demonstrate multiplexed detection of food pathogens through loop-mediated isothermal amplification on a silicon oxide chip. To our knowledge, this is the first report of miniaturized isothermal nucleic acid amplification on integrated circuit materials. Inhibition of nucleic acid amplification by adsorption of the polymerase by silicon oxide is countered with silane passivation techniques. Primers for amplification of specific infectious agents are dehydrated in silicon oxide wells, enabling multiplexed screening of virulence genes of Listeria monocytogenes, Escherichia coli, and Salmonella. Droplets of 30 nL with reagents for nucleic acid amplification and lysate of suspected pathogens are arrayed on micro-machined wells with an automated microinjection system. We show that dehydrated primers resuspend when other reagents are microinjected, and specifically amplify the targeted gene. Results of characterization experiments quantifying detection limit, specificity and robustness are also presented.
Loop-mediated isothermal amplification (LAMP), Foodborne pathogens, On-chip biomolecular detection, Parallel screening, Point-of-care (POC)
University of Illinois - Urbana-Champaign
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University of Illinois at Urbana-Champaign