Integrating Polymerase Chain Reaction, Valving, and Electrophoresis in a Plastic Device for Bacterial Detection

Abstract

An integrated plastic microfluidic device was designed and\nfabricated for bacterial detection and identification. The\ndevice, made from poly(cyclic olefin) with integrated\ngraphite ink electrodes and photopatterned gel domains,\naccomplishes DNA amplification, microfluidic valving,\nsample injection, on-column labeling, and separation.\nPolymerase chain reaction (PCR) is conducted in a\nchannel reactor containing a volume as small as 29 nL;\nthermal cycling utilizes screen-printed graphite ink resistors. In situ gel polymerization was employed to form local\nmicrofluidic valves that minimize convective flow of the\nPCR mixture into other regions. After PCR, amplicons\n(products) are electrokinetically injected through the gel\nvalve, followed by on-chip electrophoretic separation.\nAn intercalating dye is admixed to label the amplicons;\nthey are detected using laser-induced fluorescence. Two\nmodel bacteria, <i>Escherichia coli</i> O157 and <i>Salmonella\ntyphimurium</i>, were chosen to demonstrate bacterial\ndetection and identification based on amplification of\nseveral of their unique DNA sequences. The limit of\ndetection is about six copies of target DNA.