DataSheet_1_Development and evaluation of low-volume tests to detect and characterize antibodies to SARS-CoV-2.pdf

Low-volume antibody assays can be used to track SARS-CoV-2 infection rates in settings where active testing for virus is limited and remote sampling is optimal. We developed 12 ELISAs detecting total or antibody isotypes to SARS-CoV-2 nucleocapsid, spike protein or its receptor binding domain (RBD), 3 anti-RBD isotype specific luciferase immunoprecipitation system (LIPS) assays and a novel Spike-RBD bridging LIPS total-antibody assay. We utilized pre-pandemic (n=984) and confirmed/suspected recent COVID-19 sera taken pre-vaccination rollout in 2020 (n=269). Assays measuring total antibody discriminated best between pre-pandemic and COVID-19 sera and were selected for diagnostic evaluation. In the blind evaluation, two of these assays (Spike Pan ELISA and Spike-RBD Bridging LIPS assay) demonstrated >97% specificity and >92% sensitivity for samples from COVID-19 patients taken >21 days post symptom onset or PCR test. These assays offered better sensitivity for the detection of COVID-19 cases than a commercial assay which requires 100-fold larger serum volumes. This study demonstrates that low-volume in-house antibody assays can provide good diagnostic performance, and highlights the importance of using well-characterized samples and controls for all stages of assay development and evaluation. These cost-effective assays may be particularly useful for seroprevalence studies in low and middle-income countries.

低量级抗体检测技术在病毒检测受限且远程采样为优的条件下，可用来追踪 SARS-CoV-2 感染率。本研究开发了12种酶联免疫吸附测定（ELISA）用于检测 SARS-CoV-2 核衣壳、刺突蛋白或其受体结合域（RBD）的总抗体或抗体亚型，以及3种针对抗-RBD 亚型的荧光素酶免疫沉淀系统（LIPS）检测和一种新型的刺突-RBD 交联 LIPS 总抗体检测。我们利用了流感大流行前（n=984）和2020年疫苗接种推广前确诊/疑似 COVID-19 的血清样本（n=269）。测量总抗体的检测在流感大流行前和 COVID-19 血清样本之间具有最佳区分度，并被选用于诊断评估。在盲法评估中，其中两种检测（刺突全蛋白 ELISA 和刺突-RBD 交联 LIPS 检测）在症状出现或 PCR 检测后超过21天的 COVID-19 患者样本中显示出超过97%的特异性和超过92%的灵敏度。这些检测在检测 COVID-19 案例时提供的灵敏度优于需要100倍更大血清量的商业检测。本研究证实，低量级院内抗体检测能够提供良好的诊断性能，并突出了在检测开发的各个阶段使用特征明确的样本和对照的重要性。这些经济实惠的检测可能在低收入和中等收入国家的血清流行病学研究尤其有用。