Performance of nanoliter-sized droplet-based microfluidic PCR

被引:49
作者
Wang, Fang [1 ]
Burns, Mark A. [1 ,2 ]
机构
[1] Univ Michigan, Dept Chem Engn, Ann Arbor, MI 48109 USA
[2] Univ Michigan, Dept Biomed Engn, Ann Arbor, MI 48109 USA
基金
美国国家卫生研究院;
关键词
Real-time PCR; Droplet; Microfluidics; Microdevice; ELECTROWETTING-BASED ACTUATION; POLYMERASE-CHAIN-REACTION; ON-A-CHIP; PICOLITER DROPLETS; SINGLE CELLS; REAL-TIME; PROTEIN EXPRESSION; LIQUID DROPLETS; EARLY-DIAGNOSIS; DEVICE;
D O I
10.1007/s10544-009-9324-6
中图分类号
R318 [生物医学工程];
学科分类号
0831 ;
摘要
A microfluidic device was used to characterize PCR in aqueous-in-oil droplets for potential point-of-care applications. Droplets with a volume range of 5-250 nL can be formed on-chip reproducibly, and PCR in the droplets shows amplification efficiencies comparable to benchtop reactions with no evaporation loss. A higher polymerase concentration is required in the reaction droplet while the optimal Magnesium ion concentration is the same for both on-chip and benchtop systems. The optimal hold time is 9 s and 30 s for denaturation and annealing/extension in thermal cycling, respectively. With the optimized cycling parameters, the total reaction time is reduced to half of that required for benchtop PCR. For the droplets containing the same quantity of template DNA, the PCR yield is approximately the same with either fixed droplet size or fixed template DNA concentration. The droplet-based PCR can be monitored in real time with FRET probes, and provide amplification with a cycle threshold of similar to 10 cycles earlier than the benchtop instruments.
引用
收藏
页码:1071 / 1080
页数:10
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