ISGlobal, Hospital Clínic, Universitat de Barcelona, Barcelona, Catalonia, Spain
Spanish Consortium for Research in Epidemiology and Public Health (CIBERESP), Madrid, Spain.
J Clin Microbiol. 2021 Jan 21;59(2). doi: 10.1128/JCM.01731-20.
Reliable serological tests are required to determine the prevalence of antibodies against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and to characterize immunity to the disease in order to address key knowledge gaps in the coronavirus disease 2019 (COVID-19) pandemic. Quantitative suspension array technology (qSAT) assays based on the xMAP Luminex platform overcome the limitations of rapid diagnostic tests and enzyme-linked immunosorbent assays (ELISAs) with their higher precision, dynamic range, throughput, miniaturization, cost-efficiency, and multiplexing capacity. We developed three qSAT assays for IgM, IgA, and IgG against a panel of eight SARS-CoV-2 antigens, including spike protein (S), nucleocapsid protein (N), and membrane protein (M) constructs. The assays were optimized to minimize the processing time and maximize the signal-to-noise ratio. We evaluated their performances using 128 prepandemic plasma samples (negative controls) and 104 plasma samples from individuals with SARS-CoV-2 diagnosis (positive controls), of whom 5 were asymptomatic, 51 had mild symptoms, and 48 were hospitalized. Preexisting IgG antibodies recognizing N, M, and S proteins were detected in negative controls, which is suggestive of cross-reactivity to common-cold coronaviruses. The best-performing antibody/antigen signatures had specificities of 100% and sensitivities of 95.78% at ≥14 days and 95.65% at ≥21 days since the onset of symptoms, with areas under the curve (AUCs) of 0.977 and 0.999, respectively. Combining multiple markers as assessed by qSAT assays has the highest efficiency, breadth, and versatility to accurately detect low-level antibody responses for obtaining reliable data on the prevalence of exposure to novel pathogens in a population. Our assays will allow gaining insights into antibody correlates of immunity and their kinetics, required for vaccine development to combat the COVID-19 pandemic.
需要可靠的血清学检测来确定针对严重急性呼吸综合征冠状病毒 2（SARS-CoV-2）的抗体流行率，并描述针对该疾病的免疫情况，以解决 2019 年冠状病毒病（COVID-19）大流行中的关键知识空白。基于 xMAP Luminex 平台的定量悬浮阵列技术（qSAT）检测克服了快速诊断检测和酶联免疫吸附测定（ELISA）的局限性，具有更高的精度、动态范围、通量、小型化、成本效益和多重检测能力。我们开发了三种 qSAT 检测方法，用于检测针对一组 8 种 SARS-CoV-2 抗原（包括刺突蛋白（S）、核衣壳蛋白（N）和膜蛋白（M）构建体）的 IgM、IgA 和 IgG。这些检测方法经过优化，以最大程度地减少处理时间并最大程度地提高信噪比。我们使用 128 份大流行前血浆样本（阴性对照）和 104 份来自 SARS-CoV-2 诊断个体的血浆样本（阳性对照）评估了它们的性能，其中 5 例为无症状，51 例为轻症，48 例住院。在阴性对照中检测到针对 N、M 和 S 蛋白的预先存在的 IgG 抗体，这表明与普通感冒冠状病毒存在交叉反应。表现最佳的抗体/抗原特征在症状出现后≥14 天和≥21 天的特异性分别为 100%和 95.78%，敏感性分别为 95.65%，曲线下面积（AUC）分别为 0.977 和 0.999。通过 qSAT 检测评估的多种标志物组合具有最高的效率、广度和多功能性，可准确检测低水平的抗体反应，从而获得人群中对新型病原体暴露的可靠数据。我们的检测方法将有助于深入了解免疫的抗体相关性及其动力学，这是开发针对 COVID-19 大流行的疫苗所必需的。
