Guangdong Key Laboratory for Biomedical Measurements and Ultrasound Imaging, School of Biomedical Engineering, Health Science Center, Shenzhen University, Shenzhen, Guangdong 518060, People's Republic of China.
Anal Chem. 2022 Mar 15;94(10):4135-4140. doi: 10.1021/acs.analchem.2c00008. Epub 2022 Mar 2.
Rapid and high-throughput screening is critical to control the COVID-19 pandemic. Recombinase polymerase amplification (RPA) with highly accessible and sensitive nucleic acid amplification has been widely used for point-of-care infection diagnosis. Here, we report an integrated microdroplet array platform composed of an ultrasonic unit and minipillar array to enhance the RPA for ultrafast, high-sensitivity, and high-throughput detection of SARS-CoV-2. On such a platform, the independent microvolume reactions on individual minipillars greatly decrease the consumption of reagents. The microstreaming driven by ultrasound creates on-demand contactless microagitation in the microdroplets and promotes the interaction between RPA components, thus greatly accelerating the amplification. In the presence of microstreaming, the detection time is 6-12 min, which is 38.8-59.3% shorter than that of controls without microstreaming, and the end-point fluorescence intensity also increased 1.3-1.7 times. Furthermore, the microagitation-enhanced RPA also exhibits a lower detection limit (0.42 copy/μL) for SARS-CoV-2 in comparison to the controls. This integrated microdroplet array detection platform is expected to meet the needs for high-throughput nucleic acid testing (NAT) to improve the containment of viral transmission during the epidemic, as well as provide a potential platform for the timely detection of other pathogens or viruses.
快速和高通量筛选对于控制 COVID-19 大流行至关重要。具有高度可及性和敏感性的核酸扩增的重组酶聚合酶扩增（RPA）已广泛用于即时护理感染诊断。在这里，我们报告了一个由超声单元和微柱阵列组成的集成微滴阵列平台，用于增强 RPA，以实现 SARS-CoV-2 的超快、高灵敏度和高通量检测。在这样的平台上，单个微柱上的独立微体积反应大大减少了试剂的消耗。超声驱动的微流在微滴中产生按需非接触式微搅拌，并促进 RPA 组件之间的相互作用，从而大大加速了扩增。在存在微流的情况下，检测时间为 6-12 分钟，比没有微流的对照物短 38.8-59.3%，终点荧光强度也增加了 1.3-1.7 倍。此外，与对照相比，微搅拌增强的 RPA 对 SARS-CoV-2 的检测下限（0.42 拷贝/μL）也更低。这种集成的微滴阵列检测平台有望满足高通量核酸检测（NAT）的需求，以改善疫情期间病毒传播的遏制，并为其他病原体或病毒的及时检测提供潜在平台。
