Rapid and accurate diagnosis of individuals with SARS-CoV-2 infection is an effective way to prevent and control the spread of COVID-19. Although the detection of SARS‐CoV‐2 viral RNA by RT‐qPCR is the gold standard for COVID-19 testing, the use of antigen-detecting rapid diagnostic tests (Ag-RDTs) is emerging as a complementary surveillance tool as Omicron case numbers skyrocket worldwide. However, the results from Ag-RDTs are less accurate in individuals with low viral loads. To develop a highly sensitive and accurate Ag-RDT, 90 monoclonal antibodies were raised from guinea pigs immunized with SARS CoV-2 nucleocapsid protein (CoV-2-NP). By applying a capture antibody recognizing the structural epitope of the N-terminal domain of CoV-2-NP and a detection antibody recognizing the C-terminal tail of CoV-2-NP to an automated chemiluminescence flow-through membrane immunoassay device, we developed a novel Ag-RDT, CoV-2-POCube. The CoV-2-POCube exclusively recognizes CoV-2-NP variants but not the nucleocapsid proteins of other human coronaviruses. The CoV-2-POCube achieved a limit of detection sensitivity of 0.20 ~ 0.66 pg/mL of CoV-2-NPs, demonstrating more than 100 times greater sensitivity than commercially available SARS-CoV-2 Ag-RDTs. CoV-2-POCube has high analytical sensitivity and can detect SARS-CoV-2 variants in 15 min without observing the high-dose hook effect, thus meeting the need for early SARS-CoV-2 diagnosis with lower viral load. CoV-2-POCube is a promising alternative to currently available diagnostic devices for faster clinical decision making in individuals with suspected COVID-19 in resource-limited settings.

中文翻译：

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将新开发的单克隆抗体应用于自动化化学发光流通膜免疫测定装置，开发高灵敏度、定量的 SARS-CoV-2 快速抗原检测

快速、准确地诊断 SARS-CoV-2 感染者是预防和控制 COVID-19 传播的有效方法。尽管通过 RT-qPCR 检测 SARS-CoV-2 病毒 RNA 是 COVID-19 检测的黄金标准，但随着 Omicron 病例数的增加，抗原检测快速诊断检测 (Ag-RDT) 的使用正在成为一种补充监测工具在全球范围内飙升。然而，对于病毒载量较低的个体，Ag-RDT 的结果不太准确。为了开发高度灵敏和准确的 Ag-RDT，从用 SARS CoV-2 核衣壳蛋白 (CoV-2-NP) 免疫的豚鼠中提取了 90 种单克隆抗体。通过将识别 CoV-2-NP N 端结构域结构表位的捕获抗体和识别 CoV-2-NP C 端尾部的检测抗体应用于自动化化学发光流通膜免疫分析装置，我们开发了一种新型 Ag-RDT，CoV-2-POCube。CoV-2-POCube 专门识别 CoV-2-NP 变体，但不识别其他人类冠状病毒的核衣壳蛋白。CoV-2-POCube 的 CoV-2-NP 检测灵敏度极限为 0.20 ~ 0.66 pg/mL，比市售 SARS-CoV-2 Ag-RDT 的灵敏度高出 100 倍以上。CoV-2-POCube具有较高的分析灵敏度，可在15分钟内检测出SARS-CoV-2变异体，且不会观察到高剂量钩状效应，从而满足较低病毒载量的早期SARS-CoV-2诊断的需求。CoV-2-POCube 是当前可用诊断设备的一种有前景的替代品，可在资源有限的环境中更快地对疑似 COVID-19 患者做出临床决策。