Global Health Labs, Bellevue, Washington 98007, United States.
Global Good, Intellectual Ventures, Bellevue, Washington 98007, United States.
Anal Chem. 2021 Mar 9;93(9):4160-4165. doi: 10.1021/acs.analchem.0c03918. Epub 2021 Feb 25.
The rapid onset of the global COVID-19 pandemic has led to challenges for accurately diagnosing the disease, including supply shortages for sample collection, preservation, and purification. Currently, most diagnostic tests require RNA extraction and detection by RT-PCR; however, extraction is expensive and time-consuming and requires technical expertise. With these challenges in mind, we report extraction-free, multiplexed amplification of SARS-CoV-2 RNA from 246 clinical samples, resulting in 86% sensitivity and 100% specificity. The multiplex RT-PCR uses the CDC singleplex targets and has an LoD of 2 c/μL. We also report on amplification using a range of master mixes in different transport media. This work can help guide which combinations of reagents will enable accurate results when availability of supplies changes throughout the pandemic. Implementing these methods can reduce complexity and cost, minimize reagent usage, expedite time to results, and increase testing capacity.
全球 COVID-19 大流行的迅速爆发给疾病的准确诊断带来了挑战，包括样本采集、保存和纯化方面的供应短缺。目前，大多数诊断测试都需要 RNA 提取和 RT-PCR 检测；然而，提取既昂贵又耗时，并且需要技术专业知识。考虑到这些挑战，我们报告了从 246 个临床样本中无提取、多重扩增 SARS-CoV-2 RNA，灵敏度为 86%，特异性为 100%。多重 RT-PCR 使用 CDC 单重靶标，检测限为 2 c/μL。我们还报告了使用不同运输介质中的一系列主混合物进行扩增的情况。这项工作可以帮助指导在整个大流行期间供应情况发生变化时，哪些试剂组合将能够获得准确的结果。实施这些方法可以降低复杂性和成本，最小化试剂使用量，加快结果产生时间，并提高检测能力。
