Abstract
Polymerase chain reaction (PCR) is a powerful tool for nucleic acid amplification and quantification. However, long thermocycling time is a major limitation of the commercial PCR devices in the point-of-care (POC). Herein, we have developed a rapid droplet-based photonic PCR (dpPCR) system, including a gold (Au) nanofilm-based microfluidic chip and a plasmonic photothermal cycler. The chip is fabricated by adding mineral oil to uncured polydimethylsiloxane (PDMS) to suppress droplet evaporation in PDMS microfluidic chips during PCR thermocycling. A PDMS to gold bonding technique using a double-sided adhesive tape is applied to enhance the bonding strength between the oil-added PDMS and the gold nanofilm. Moreover, the gold nanofilm excited by two light-emitting diodes (LEDs) from the top and bottom sides of the chip provides fast heating of the PCR sample to 230 °C within 100 s. Such a design enables 30 thermal cycles from 60 to 95 °C within 13 min with the average heating and cooling rates of 7.37 ± 0.27 °C/s and 1.91 ± 0.03 °C/s, respectively. The experimental results demonstrate successful PCR amplification of the alcohol oxidase (AOX) gene using the rapid plasmonic photothermal cycler and exhibit the great performance of the microfluidic chip for droplet-based PCR.
Highlights
Polymerase chain reaction (PCR) is a powerful tool for nucleic acid amplification and quantification
We describe a simple, rapid, inexpensive, and reliable system for droplet-based photonic PCR analysis by integrating the functions of microdroplet generation, plasmonic photothermal heating of Au nanofilms (AuNFs), and PDMS microfluidic chip
We present a rapid plasmonic photothermal cycler consisting of a thin Au film and two blue light-emitting diodes (LEDs) for fast droplet-based PCR, providing a heating rate of 7.37 ± 0.27 °C/s and a cooling rate of 1.91 ± 0.03 °C/s
Summary
Polymerase chain reaction (PCR) is a powerful tool for nucleic acid amplification and quantification. Conventional PCR systems, which utilize a Peltier-based thermal cycler, are capable of dramatically amplifying target nucleic acids They require a long thermocycling time since the heat transfer between the heating block and the plastic PCR tube is slow[43]. AuNFs may be regarded as a better alternative for nucleic acid amplification than AuNPs. More recently, the photonic thermocycling method, based on the photothermal heating of AuNFs using LED irradiation, has received ever-increasing attention all over the world. The photonic thermocycling method, based on the photothermal heating of AuNFs using LED irradiation, has received ever-increasing attention all over the world The advantages such as low power consumption, rapid temperature ramping capability, and low-cost make this method more beneficial for POC testing[51]
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