Beck Sungjun, Nakajima Rie, Jasinskas Algis, Abram Timothy J, Kim Sun Jin, Bigdeli Nader, Tifrea Delia F, Hernandez-Davies Jenny, Huw Davies D, Hedde Per Niklas, Felgner Philip L, Zhao Weian
Department of Biological Chemistry, University of California, Irvine, CA 92697, USA.
Department of Physiology and Biophysics, University of California, Irvine, CA 92697, USA.
Biomedicines. 2022 Sep 9;10(9):2238. doi: 10.3390/biomedicines10092238.
High-throughput and rapid screening testing is highly desirable to effectively combat the rapidly evolving COVID-19 pandemic co-presents with influenza and seasonal common cold epidemics. Here, we present a general workflow for iterative development and validation of an antibody-based microarray assay for the detection of a respiratory viral panel: (a) antibody screening to quickly identify optimal reagents and assay conditions, (b) immunofluorescence assay design including signal amplification for low viral titers, (c) assay characterization with recombinant proteins, inactivated viral samples and clinical samples, and (d) multiplexing to detect a panel of common respiratory viruses. Using RT-PCR-confirmed SARS-CoV-2 positive and negative pharyngeal swab samples, we demonstrated that the antibody microarray assay exhibited a clinical sensitivity and specificity of 77.2% and 100%, respectively, which are comparable to existing FDA-authorized antigen tests. Moreover, the microarray assay is correlated with RT-PCR cycle threshold (Ct) values and is particularly effective in identifying high viral titers. The multiplexed assay can selectively detect SARS-CoV-2 and influenza virus, which can be used to discriminate these viral infections that share similar symptoms. Such protein microarray technology is amenable for scale-up and automation and can be broadly applied as a both diagnostic and research tool.
高通量和快速筛查检测对于有效应对迅速演变的新冠疫情以及同时出现的流感和季节性普通感冒疫情非常必要。在此，我们展示了一种用于迭代开发和验证基于抗体的微阵列检测方法以检测呼吸道病毒组的通用工作流程：(a) 抗体筛选以快速确定最佳试剂和检测条件，(b) 免疫荧光检测设计，包括针对低病毒滴度的信号放大，(c) 用重组蛋白、灭活病毒样本和临床样本进行检测表征，以及 (d) 多重检测以检测一组常见呼吸道病毒。使用经逆转录聚合酶链反应（RT-PCR）确认的新冠病毒阳性和阴性咽拭子样本，我们证明该抗体微阵列检测方法的临床敏感性和特异性分别为77.2%和100%，与现有的美国食品药品监督管理局（FDA）授权的抗原检测相当。此外，该微阵列检测方法与RT-PCR循环阈值（Ct）值相关，并且在识别高病毒滴度方面特别有效。这种多重检测方法可以选择性地检测新冠病毒和流感病毒，可用于区分这些具有相似症状的病毒感染。这种蛋白质微阵列技术适合扩大规模和自动化，并且可以广泛用作诊断和研究工具。
