Rapid visual detection of hepatitis C virus using a reverse transcription loop-mediated isothermal amplification assay.
Rapid visual detection of hepatitis C virus using a reverse transcription loop-mediated isothermal amplification assay.
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使用逆转录环介导的等温扩增测定法快速目视检测丙型肝炎病毒。
DOI：
10.1016/j.ijid.2020.10.082
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发表时间：
2021-01
期刊：
影响因子：
--
通讯作者：
Supparatpinyo K
中科院分区：
文献类型：
--
作者：
Hongjaisee S;Doungjinda N;Khamduang W;Carraway TS;Wipasa J;Debes JD;Supparatpinyo K
目的：建立一种封闭单管逆转录环介导等温扩增（RT-LAMP）检测丙型肝炎病毒（HCV）的方法。从200例HCV感染患者中收集血浆样品。HCV-RNA采用一步法RT-LAMP检测，反应温度65 °C，反应时间60 min，扩增产物采用羟萘酚蓝（HNB）目视法和凝胶电泳法检测。检测了针对其他病毒的特异性。使用提取的RNA的连续稀释液测定灵敏度。RT-LAMP检测HCV-RNA基因1型检出率为97.5%，3型为91.1%，6型为100%。肉眼可观察到颜色变化。该测定法证明临床灵敏度为95.5%，特异性为100%，并且与其他病毒无交叉反应性（即，肝炎B病毒，HIV）。HCV基因型1a和6的检测限低至10 ng/反应，而基因型3a的检测限为100 ng/反应。在检测的临床样本中，当病毒载量为5.00 log 10 IU/mL时，检测试剂盒显示100%检出阈值。这项研究表明，使用RT-LAMP检测HCV的一个简单，快速，具有成本效益的方式，这将是有用的，在资源有限的设置，以允许个人在需要HCV治疗的识别。
The aim was to develop a reverse transcription loop-mediated isothermal amplification (RT-LAMP) assay for the detection of hepatitis C virus (HCV) in a single closed tube. Plasma samples were collected from 200 HCV-infected patients. HCV-RNA was detected by one-step RT-LAMP processed at 65 °C for 60 min. The amplified products were detected by hydroxynaphthol blue (HNB)-dependent visual method and gel electrophoresis. Specificity was tested against other viruses. Sensitivity was determined using serial dilutions of extracted RNA. The RT-LAMP assay detected 97.5% of HCV-RNA genotype 1, 91.1% of genotype 3, and 100% of genotype 6. The color change was evidenced with the naked eye. The assay demonstrated a clinical sensitivity of 95.5% and specificity of 100%, as well as no cross-reactivity with other viruses (i.e., hepatitis B virus, HIV). The limit of detection was as low as 10 ng per reaction for HCV genotypes 1a and 6, while it was 100 ng for genotype 3a. The assay showed a 100% detection threshold at a viral load of 5.00 log10 IU/mL in the clinical samples tested. This study demonstrated the use of an RT-LAMP assay for the detection of HCV in a simple, rapid, and cost-effective manner, which will be useful in resource-limited settings to allow the identification of individuals in need of HCV treatment.
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影响因子：
9.4
作者：
Parida, M;Posadas, G;Morita, K
通讯作者：
Morita, K
DOI：
10.1016/j.diagmicrobio.2008.04.001
发表时间：
2008-08-01
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作者：
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