Real-time PCR detection of Listeria monocytogenes using an integrated microfluidics platform

A miniaturized, fully automated, PCR-based detection system has been developed for the rapid detection of bacterial pathogens. Monolithic DNA purification/real-time PCR silicon chips were fabricated and tested for their ability to purify and detect the pathogeneic bacterium Listeria monocytogenes. Using silica-coated microstructures, nucleic acids could be selectively bound, washed and eluted for subsequent real-time PCR. These microstructures were included in an integrated detection microchip containing two distinct regions, one for DNA purification and one for real-time PCR. Using an automated detection platform with integrated microprocessor, pumps, valves, thermocycler and fluorescence detection modules, microchips were used to purify and detect bacterial DNA by real-time PCR amplification using SYBR Green fluorescent dye. Between 10(4) and 10(7) L. monocytogenes cells could be detected using this system with an average turnaround time of 45 min. (c) 2004 Elsevier B.V. All rights reserved.