Glass-polytetrafluoroethylene-glass based sandwich microdevice for continuous-flow polymerase chain reaction and its application for fast identification of foodborne pathogens

Abstract

A uniquely fabricated microdevice with a glass-polytetrafluoroethylene (PTFE)-glass (GPG) sandwich configuration was developed for the fast identification of foodborne pathogens by using continuous-flow polymerase chain reaction (PCR) followed by on-site fluorescence detection of PCR products. A PTFE tube was simply folded into a serpentine configuration and sandwiched between two glass slides to maintain a 2D planar structure. By using a PTFE tube, the microchannel has the advantage of seamlessness without the need for photolithography methods, and is less expensive and less time-consuming to fabricate than alternative devices. The non-porous and gas-impermeable PTFE tube ensured stable sample flow inside the microchannel. PCR conditions were optimized and evaluated by examining the effects of number of cycles and initial concentration of bacteria introduced. On-site detection of PCR products was performed using nucleic acid dye SYBR Green I. Using the GPG microdevice, Escherichia coli O157:H7 and Salmonella spp., two well-known foodborne pathogens, were amplified within 20 min. Besides fast reaction, the simply fabricated microdevice demonstrated its potential as a portable device applicable for public healthcare diagnostics.