Rapid and sensitive SARS-CoV-2 detection using a homogeneous fluorescent immunosensor Quenchbody with crowding agents
Rapid and sensitive SARS-CoV-2 detection using a homogeneous fluorescent immunosensor Quenchbody with crowding agents
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DOI：
10.1039/d2an01051h
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发表时间：
2022-09-30
期刊：
影响因子：
4.2
通讯作者：
Ueda, Hiroshi
中科院分区：
文献类型：
--
作者：
Zhu, Bo;Nosaka, Nobuyuki;Ueda, Hiroshi
在持续的COVID-19大流行期间，公众广泛使用SARS-CoV-2抗原检测，这表明了同质免疫传感器相关技术的社会影响。本研究利用PM Q探针技术和Quenchbody技术，研制了一种基于人源抗SARS冠状病毒N蛋白抗体的均相免疫传感器。首次揭示了拥挤剂在提高双标记淬灭体性能方面的作用，并讨论了这种改进背后的可能机制。5%聚乙二醇6000显著提高了SARS-CoV-2 Quenchbodies的响应速度和灵敏度。在孵育15 min内，重组N蛋白的计算检测限为191 pM（9 ng mL（-1）），比不添加拥挤剂的测定低9- 10倍。我们还通过使用紧凑型管荧光计测量COVID-19阳性患者的标本，在即时检测中验证了所开发的免疫传感器。简而言之，这项工作显示了Quenchbody均相免疫传感器作为快速和具有成本效益的工具的可行性，用于在COVID-19或其他新兴传染病的大规模监测和流行病学研究中对拭子样本进行诊断和高通量分析。
Antigen tests for SARS-CoV-2 are widely used by the public during the ongoing COVID-19 pandemic, which demonstrates the societal impact of homogeneous immunosensor-related technologies. In this study, we used the PM Q-probe and Quenchbody technologies to develop a SARS-CoV-2 nucleocapsid protein (N protein) homogeneous immunosensor based on a human anti-N protein antibody. For the first time, we uncovered the crowding agent's role in improving the performance of the double-labeled Quenchbody, and the possible mechanisms behind this improvement are discussed. The 5% polyethylene glycol 6000 significantly improved both the response speed and sensitivity of SARS-CoV-2 Quenchbodies. The calculated limit of detection for recombinant N protein was 191 pM (9 ng mL(-1)) within 15 min of incubation, which was 9- to 10-fold lower than the assay without adding crowding agent. We also validated the developed immunosensor in a point-of-care test by measuring specimens from COVID-19-positive patients using a compact tube fluorometer. In brief, this work shows the feasibility of Quenchbody homogeneous immunosensors as rapid and cost-efficient tools for the diagnosis and high-throughput analysis of swab samples in large-scale monitoring and epidemiological studies of COVID-19 or other emerging infectious diseases.