title: Rapid and highly sensitive detection by a real-time polymerase chain reaction using a chip coated with its reagents.
abstract: On-site detection by flow-through polymerase chain reaction (PCR) microfluidic systems for rapid and highly sensitive analysis, are significantly desired for bioanalytical and medical research. The conventional continuous-flow PCR chips realized rapid detection, but their sensitivity was very low (10(6) to 10(8) copies μL(-1)). We improved this drawback by coating the chip with a PCR reagents mixture, and succeed to obtain a rapid and highly sensitive detection by using a segment-flow PCR system. In the present work, we developed a portable segment-flow PCR system for practical use. PCR was performed for the uid A gene in E. coli. By real-time segment-flow PCR using coated chips, we realized rapid detection in 8 min and a high sensitivity of 4 cells μL(-1). The sensitivity by the segment-flow PCR chip was the same as that of a conventional thermal cycler. Moreover, the detection speed of our segment-flow PCR chip was 15-times as rapid as that of the conventional thermal cycler.
authors.affiliation: Health Research Institute, National Institute of Advanced Industrial Science and Technology (AIST).
authors.forename: Shunsuke
authors.initials: S
authors.lastname: Furutani
authors.forename: Nahoko
authors.initials: N
authors.lastname: Naruishi
authors.forename: Masato
authors.initials: M
authors.lastname: Saito
authors.forename: Eiichi
authors.initials: E
authors.lastname: Tamiya
authors.forename: Yusuke
authors.initials: Y
authors.lastname: Fuchiwaki
authors.forename: Hidenori
authors.initials: H
authors.lastname: Nagai
mesh_terms: D004269:DNA, Bacterial / Q000235:genetics / Q000302:isolation & purification; D004867:Equipment Design; D004926:Escherichia coli / Q000235:genetics / Q000302:isolation & purification; D007202:Indicators and Reagents; D046210:Microfluidic Analytical Techniques / Q000295:instrumentation* / Q000379:methods*; D060888:Real-Time Polymerase Chain Reaction / Q000295:instrumentation* / Q000379:methods*; D012680:Sensitivity and Specificity; D013997:Time Factors
chemical_list: D004269:DNA, Bacterial; D007202:Indicators and Reagents