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基于纳米粒子的扩增 DNA 可视化检测在丙型肝炎病毒诊断中的应用。

Nanoparticle-Based Visual Detection of Amplified DNA for Diagnosis of Hepatitis C Virus.

机构信息

Department of Laboratory Medicine, Ewha Womans University College of Medicine, Seoul 07985, Korea.

Department of Laboratory Medicine, Asan Medical Center, University of Ulsan College of Medicine, Seoul 05505, Korea.

出版信息

Biosensors (Basel). 2022 Sep 9;12(9):744. doi: 10.3390/bios12090744.

Abstract

Rapid, simple, and inexpensive diagnostic point-of-care tests (POCTs) are essential for controlling infectious diseases in resource-limited settings. In this study, we developed a new detection system based on nanoparticle-DNA aggregation (STat aggregation of tagged DNA, STAT-DNA) to yield a visual change that can be easily detected by the naked eye. This simplified optical detection system was applied to detect hepatitis C virus (HCV). Reverse transcription-polymerase chain reaction (RT-PCR) was performed using primers labeled with biotin and digoxigenin. Streptavidin-coated magnetic particles (1 μm) and anti-digoxigenin antibody-coated polystyrene particles (250-350 nm) were added to form aggregates. The limit of detection (LoD) and analytical specificity were analyzed. The STAT-DNA results were compared with those of the standard real-time PCR assay using serum samples from 54 patients with hepatitis C. We achieved visualization of amplified DNA with the naked eye by adding nanoparticles to the PCR mixture without employing centrifugal force, probe addition, incubation, or dilution. The LoD of STAT-DNA was at least 10 IU/mL. STAT-DNA did not show cross-reactivity with eight viral pathogens. The detection using STAT-DNA was consistent with that using standard real-time PCR.

摘要

在资源有限的环境中,快速、简便且廉价的即时诊断检测(POCT)对于传染病的控制至关重要。在这项研究中,我们开发了一种基于纳米粒子-DNA 聚集(标记 DNA 的 STAT 聚集,STAT-DNA)的新型检测系统,可产生肉眼可轻松检测到的视觉变化。该简化的光学检测系统被应用于检测丙型肝炎病毒(HCV)。使用带有生物素和地高辛标记的引物进行逆转录-聚合酶链反应(RT-PCR)。添加链霉亲和素包被的磁性颗粒(1 μm)和抗地高辛抗体包被的聚苯乙烯颗粒(250-350nm)以形成聚集体。分析了检测限(LoD)和分析特异性。使用来自 54 名丙型肝炎患者的血清样本比较了 STAT-DNA 结果与标准实时 PCR 检测结果。通过将纳米粒子添加到 PCR 混合物中,而无需离心、探针添加、孵育或稀释,我们实现了肉眼可见的扩增 DNA。STAT-DNA 的 LoD 至少为 10IU/mL。STAT-DNA 与八种病毒病原体无交叉反应。STAT-DNA 的检测结果与标准实时 PCR 检测结果一致。

https://cdn.ncbi.nlm.nih.gov/pmc/blobs/1fe8/9496050/02f17eddd4d9/biosensors-12-00744-g001.jpg

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