CRISPR-Cas13a 与手机显微镜无扩增检测 SARS-CoV-2。
Amplification-free detection of SARS-CoV-2 with CRISPR-Cas13a and mobile phone microscopy.
机构信息
J. David Gladstone Institutes, San Francisco, CA 94158, USA; Medical Scientist Training Program, University of California, San Francisco, San Francisco, CA 94143, USA; Biomedical Sciences Graduate Program, University of California, San Francisco, San Francisco, CA 94143, USA; Department of Medicine, University of California, San Francisco, San Francisco, CA 94143, USA.
Department of Bioengineering, University of California, Berkeley, Berkeley, CA 94720, USA.
出版信息
Cell. 2021 Jan 21;184(2):323-333.e9. doi: 10.1016/j.cell.2020.12.001. Epub 2020 Dec 4.
The December 2019 outbreak of a novel respiratory virus, SARS-CoV-2, has become an ongoing global pandemic due in part to the challenge of identifying symptomatic, asymptomatic, and pre-symptomatic carriers of the virus. CRISPR diagnostics can augment gold-standard PCR-based testing if they can be made rapid, portable, and accurate. Here, we report the development of an amplification-free CRISPR-Cas13a assay for direct detection of SARS-CoV-2 from nasal swab RNA that can be read with a mobile phone microscope. The assay achieved ∼100 copies/μL sensitivity in under 30 min of measurement time and accurately detected pre-extracted RNA from a set of positive clinical samples in under 5 min. We combined crRNAs targeting SARS-CoV-2 RNA to improve sensitivity and specificity and directly quantified viral load using enzyme kinetics. Integrated with a reader device based on a mobile phone, this assay has the potential to enable rapid, low-cost, point-of-care screening for SARS-CoV-2.
2019 年 12 月，一种新型呼吸道病毒 SARS-CoV-2 的爆发成为一场持续的全球大流行，部分原因是难以识别病毒的有症状、无症状和症状前携带者。如果 CRISPR 诊断能够快速、便携且准确，它们可以增强基于聚合酶链反应（PCR）的金标准检测。在这里，我们报告了一种扩增免 PCR 的 CRISPR-Cas13a 检测方法的开发，该方法可直接从鼻拭子 RNA 中检测 SARS-CoV-2，并用手机显微镜读取结果。该检测方法在 30 分钟的测量时间内达到了约 100 拷贝/μL 的灵敏度，并在 5 分钟内准确检测了一组阳性临床样本中的预提取 RNA。我们将靶向 SARS-CoV-2 RNA 的 crRNA 组合在一起，以提高灵敏度和特异性，并直接通过酶动力学定量病毒载量。与基于手机的读取器设备集成后，该检测方法有可能实现 SARS-CoV-2 的快速、低成本、即时护理筛查。
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