College of Chemistry and Molecular Sciences, Wuhan University, Wuhan 430072, P. R. China.
Hubei Provincial Center for Disease Control and Prevention, Wuhan 430079, P. R. China.
ACS Sens. 2021 Mar 26;6(3):1086-1093. doi: 10.1021/acssensors.0c02365. Epub 2021 Mar 8.
The outbreak of COVID-19 caused a worldwide public health crisis. Large-scale population screening is an effective means to control the spread of COVID-19. Reverse transcription-polymerase chain reaction (RT-qPCR) and serology assays are the most available techniques for SARS-CoV-2 detection; however, they suffer from either less sensitivity and accuracy or low instrument accessibility for screening. To balance the sensitivity, specificity, and test availability, here, we developed enhanced colorimetry, which is termed as a magnetic pull-down-assisted colorimetric method based on the CRISPR/Cas12a system (M-CDC), for SARS-CoV-2 detection. By this method, SARS-CoV-2 RNA from synthetic sequences and cultured viruses can be detected by the naked eye based on gold nanoparticle (AuNP) probes, with a detection limit of 50 RNA copies per reaction. With CRISPR/Cas12a-assisted detection, SARS-CoV-2 can be specifically distinguished from other closely related viruses. M-CDC was further used to analyze 41 clinical samples, whose performance was 95.12%, consistent with that of an approved Clinical RT-qPCR Diagnosis kit. The developed M-CDC method is not dependent on sophisticated instruments, which makes it potentially valuable to be applied for SARS-CoV-2 screening under poor conditions.
新型冠状病毒肺炎疫情的爆发引发了全球性的公共卫生危机。大规模人群筛查是控制新型冠状病毒肺炎传播的有效手段。逆转录聚合酶链反应（RT-qPCR）和血清学检测是目前最常用的检测 SARS-CoV-2 的技术；然而，它们要么敏感性和准确性较低，要么仪器难以普及，不适合用于筛查。为了平衡敏感性、特异性和检测可用性，我们在这里开发了增强比色法，称为基于 CRISPR/Cas12a 系统的磁下拉辅助比色法（M-CDC），用于检测 SARS-CoV-2。通过该方法，基于金纳米粒子（AuNP）探针，可以通过肉眼检测到合成序列和培养病毒的 SARS-CoV-2 RNA，检测限为每个反应 50 个 RNA 拷贝。通过 CRISPR/Cas12a 辅助检测，可以特异性地区分 SARS-CoV-2 与其他密切相关的病毒。M-CDC 进一步用于分析 41 个临床样本，其性能为 95.12%，与经批准的临床 RT-qPCR 诊断试剂盒相当。所开发的 M-CDC 方法不依赖于复杂的仪器，使其具有在条件较差的情况下用于 SARS-CoV-2 筛查的潜在价值。
