Department of Microbiology, College of Natural Sciences, Chungbuk National University, Cheongju, Republic of Korea.
Infectious Diseases Research Center, Korea Research Institute of Bioscience and Biotechnology, Daejeon, Republic of Korea.
Sci Rep. 2019 Nov 13;9(1):16661. doi: 10.1038/s41598-019-53083-8.
Point-of-care tests (POCT) for pathogens are considered important for low-resource countries and facilities. Although lateral flow immunoassays (LFIA) have many advantages including speed and ease of use, their sensitivity is limited without specific equipment. Furthermore, their response cannot be enhanced through enzymatic reactions. Owing to these limitations, LFIAs have not yet been generally adopted as the standard protocol for in vitro analysis of infectious pathogens. We aimed to develop a novel pipetting-based immunoassay using a removable magnetic ring-coupled pipette tip. The "magnetic bead-capture antibody-targeted protein complex" was simply purified by pipetting and quantified by enzymatic colour development or using a lateral flow system. This pipetting-based immunoassay was applied to detect the nucleoprotein (NP) of the influenza A virus. Using an HRP-conjugated monoclonal antibody as a probe, the assay allowed for specific and sensitive detection. Furthermore, when this assay was applied exclusively for antigen capture in the lateral flow system, the limit of detection improved 100-fold and displayed greater sensitivity than the lateral flow system alone. Therefore, the pipetting-based immunoassay may be potentially used as a sensitive POCT to clinically detect a target antigen.
即时检测（POCT）病原体在资源匮乏的国家和医疗机构中具有重要意义。虽然侧向流动免疫分析（LFIA）具有速度快、使用方便等优点，但如果没有特定的设备，其灵敏度就会受到限制。此外，其反应也无法通过酶反应来增强。由于这些局限性，LFIA 尚未被普遍采用为感染性病原体体外分析的标准方案。本研究旨在开发一种新型基于移液的免疫分析方法，该方法使用可移动的磁环耦合移液吸头。通过移液，简单地纯化“磁珠-捕获抗体-靶蛋白复合物”，并通过酶显色或使用侧向流动系统进行定量。该基于移液的免疫分析用于检测甲型流感病毒的核蛋白（NP）。使用 HRP 标记的单克隆抗体作为探针，该检测方法具有特异性和敏感性。此外，当该检测方法仅应用于侧向流动系统中的抗原捕获时，检测限提高了 100 倍，比单独使用侧向流动系统更灵敏。因此，基于移液的免疫分析可能具有作为临床检测靶抗原的灵敏 POCT 的潜力。
