The coronavirus disease 2019 (COVID-19) pandemic caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is in urgent need of therapeutic options. High-throughput screening (HTS) offers an opportunity to rapidly identify such compounds. In this work, we have developed a homogeneous cell-based HTS system using AlphaLISA detection technology for the SARS-CoV-2 nucleocapsid protein (NP). Our assay measures both recombinant and endogenous NP from viral lysates and tissue culture supernatants (TCS) in a sandwich-based format using two monoclonal antibodies against the NP analyte. Viral NP was detected and quantified in both tissue culture supernatants and cell lysates, with large differences observed between 24 and 48 h of infection. We simulated viral infection by spiking recombinant NP into 384-well plates with live Vero-E6 cells and were able to detect the NP with high sensitivity and a large dynamic range. Antiviral agents that inhibit either viral cell entry or replication decrease the AlphaLISA NP signal. Thus, this assay can be used for high-throughput screening of small molecules and biologics in the fight against the COVID-19 pandemic.

中文翻译：

---

开发用于检测 SARS-CoV-2 核衣壳的高通量均质 AlphaLISA 药物筛选试验

由严重急性呼吸综合征冠状病毒 2 (SARS-CoV-2) 引起的 2019 年冠状病毒病 (COVID-19) 大流行迫切需要治疗选择。高通量筛选（HTS）提供了快速鉴定此类化合物的机会。在这项工作中，我们使用 AlphaLISA 检测技术开发了一种基于细胞的同质 HTS 系统，用于 SARS-CoV-2 核衣壳蛋白 (NP)。我们的测定使用两种针对 NP 分析物的单克隆抗体，以夹心形式测量来自病毒裂解物和组织培养上清液 (TCS) 的重组和内源 NP。在组织培养上清液和细胞裂解物中均检测到病毒 NP 并进行定量，在感染 24 至 48 小时之间观察到较大差异。我们通过将重组 NP 掺入含有活 Vero-E6 细胞的 384 孔板中来模拟病毒感染，并能够以高灵敏度和大动态范围检测 NP。抑制病毒细胞进入或复制的抗病毒药物会降低 AlphaLISA NP 信号。因此，该测定可用于对抗 COVID-19 大流行的小分子和生物制剂的高通量筛选。