A Practical Liquid Plug Flow-through Polymerase Chain-Reaction System Based on a Heat-Resistant Resin Chip

Abstract

Flow-through polymerase chain reaction (PCR) microfluidic systems for fast, small-volume DNA amplification on a single chip are significantly impacting medical and bioanalytical research. We have fabricated an improved, practical flow-through PCR chip by weighting a pressure-sensitive polyolefin (PSP) film onto a cyclo-olefin polymer (COP) substrate. The substrate was cut so as to produce microchannels, and was used to amplify DNA using a small moving liquid plug, in contrast to conventional continuous-flow-through PCR. Infrared (IR) imaging-based thermal analysis of the PSP film enabled the gradient and uniformity of the rapidly flowing microfluid to be accurately visualized, because the PSP film is thin, transparent and heat resistant. The liquid plug flow-through PCR, operating at 150 μl min -1 , showed approximately 55% amplification compared to a commercial PCR instrument, and required only 250 s for 40 cycles. To our knowledge, this is the fastest cycling time reported to date. In a real test sample, the liquid plug flow-through PCR could detect the presence or absence of anthrax in a sample solution in approximately 250 s. Thus, this liquid plug flow-through PCR system, based on our novel PCR chip, excelled in a practical applications compared to other devices.

Analysis of PCR Kinetics inside a Microfluidic DNA Amplification System

“…To construct a CFPCR chip employing one heater or two heaters, the extension regions or the annealing regions are generated by the thermal gradients between the high-temperature and low-temperature regions. In previous studies, the temperature distributions inside the reaction regions were demonstrated numerically and/or experimentally and utilized to ensure that the requisite temperatures for PCR were existed [ 23 , 24 ]. In addition, by the rearrangement and enlargement of the channel geometry at the extension region, the residence time during the extension step can be properly extended to complete PCR [ 25 ].…”

Portable Microfluidic System for Rapid Genetic Testing

“…In contrast, we developed segment‐flow PCR to prevent interruption of continuous flow; in particular, the sample solution is transported as a liquid plug with respect to the gas phase, thus releasing spontaneous air bubbles into the gas phase . This is equivalent to continuous‐flow PCR in that the sample solution is thermally cycled while flowing through multiple temperature zones of a serpentine microchannel.…”