Department of Dermatology, University Hospital Zurich, University of Zurich, Zurich, Switzerland.
ADMed Microbiologie, La Chaux-de-Fonds, Switzerland.
J Mol Diagn. 2021 Jul;23(7):796-804. doi: 10.1016/j.jmoldx.2021.04.009. Epub 2021 May 5.
Real-time RT-PCR remains a gold standard in the detection of various viral diseases. In the coronavirus 2019 pandemic, multiple RT-PCR-based tests were developed to screen for viral infection. As an emergency response to increasing testing demand, we established a severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) PCR diagnostics platform for which we compared different commercial and in-house RT-PCR protocols. Four commercial, one customized, and one in-house RT-PCR protocols were evaluated with 92 SARS-CoV-2-positive and 92 SARS-CoV-2-negative samples. Furthermore, economical and practical characteristics of these protocols were compared. In addition, a highly sensitive digital droplet PCR (ddPCR) method was developed, and application of RT-PCR and ddPCR methods on SARS-CoV-2 environmental samples was examined. Very low limits of detection (1 or 2 viral copies/μL), high sensitivities (93.6% to 97.8%), and high specificities (98.7% to 100%) for the tested RT-PCR protocols were found. Furthermore, the feasibility of downscaling two of the commercial protocols, which could optimize testing capacity, was demonstrated. Tested commercial and customized RT-PCR detection kits show very good and comparable sensitivity and specificity, and the kits could be further optimized for use on SARS-CoV-2 viral samples derived from human and surface swabbed samples.
实时 RT-PCR 仍然是检测各种病毒疾病的金标准。在 2019 年冠状病毒病大流行期间，开发了多种基于 RT-PCR 的检测方法来筛查病毒感染。作为对检测需求不断增加的紧急应对措施，我们建立了一种严重急性呼吸系统综合征冠状病毒 2 (SARS-CoV-2) PCR 诊断平台，我们比较了不同的商业和内部 RT-PCR 方案。我们用 92 份 SARS-CoV-2 阳性和 92 份 SARS-CoV-2 阴性样本评估了四种商业、一种定制和一种内部 RT-PCR 方案。此外，还比较了这些方案的经济实用特性。此外，还开发了一种高灵敏度的数字液滴 PCR (ddPCR) 方法，并研究了 RT-PCR 和 ddPCR 方法在 SARS-CoV-2 环境样本中的应用。所测试的 RT-PCR 方案具有非常低的检测限（1 或 2 个病毒拷贝/μL）、高灵敏度（93.6%至 97.8%）和高特异性（98.7%至 100%）。此外，还证明了两种商业方案的缩微化的可行性，这可以优化测试能力。测试的商业和定制 RT-PCR 检测试剂盒显示出非常好和可比的灵敏度和特异性，并且可以进一步优化用于检测来自人类和表面擦拭样本的 SARS-CoV-2 病毒样本。
