Rapid and Visual Detection of SARS-CoV-2 Using Multiplex Reverse Transcription Loop-Mediated Isothermal Amplification Linked With Gold Nanoparticle-Based Lateral Flow Biosensor.
Rapid and Visual Detection of SARS-CoV-2 Using Multiplex Reverse Transcription Loop-Mediated Isothermal Amplification Linked With Gold Nanoparticle-Based Lateral Flow Biosensor.
复制标题
使用多重逆转录环介导的等温扩增与基于金纳米颗粒的侧流生物传感器相结合，快速、直观地检测 SARS-CoV-2。
DOI：
10.3389/fcimb.2021.581239
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发表时间：
2021
影响因子：
5.7
通讯作者：
Dong S
中科院分区：
文献类型：
--
作者：
Chen X;Zhou Q;Li S;Yan H;Chang B;Wang Y;Dong S
严重急性呼吸综合征冠状病毒2型（SARS-CoV-2）是一种新型冠状病毒，导致2019冠状病毒病（COVID-19）在全球爆发。在缺乏合适的抗病毒药物或疫苗的情况下，开发一种简单、快速和可靠的SARS-CoV-2检测方法对于预防和控制COVID-19传播是必要的。基于SARS-CoV-2的RdRp和N基因，将一种新型的分子诊断技术--多重逆转录环介导等温扩增技术（mRT-LAMP-LFB）与基于纳米颗粒的侧流生物传感器（nanoparticle-based lateral flow biosensor）相连接，用于检测SARS-CoV-2，并利用基于纳米颗粒的侧流生物传感器分析mRT-LAMP产物。优化了mRT-LAMP-LFB扩增条件，包括目标RNA浓度、扩增温度和时间。本研究对mRT-LAMP-LFB方法的敏感性和特异性进行了检测，并将mRT-LAMP-LFB方法应用于临床标本和人工痰标本中SARS-CoV-2的检测。根据SARS冠状病毒RdRp和N基因设计的特异性引物可用于建立检测SARS冠状病毒的mRT-LAMP-LFB方法。多重RT-LAMP扩增条件在63°C下优化30分钟。从反应前准备、病毒RNA提取、RT-LAMP扩增、产物鉴定等全过程只需80 min。mRT-LAMP-LFB技术的检测限（LoD）为每个反应20个拷贝。mRT-LAMP-LFB检测的特异性为100%，与其他呼吸道病原体无交叉反应。本研究开发的mRT-LAMP-LFB技术是一种简单、快速、可靠的方法，在识别SARS-CoV-2病毒以预防和控制COVID-19疾病方面具有很高的特异性和灵敏度，特别是在世界资源有限的地区。
Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is a novel coronavirus that has caused the outbreak of coronavirus disease 2019 (COVID-19) all over the world. In the absence of appropriate antiviral drugs or vaccines, developing a simple, rapid, and reliable assay for SARS-CoV-2 is necessary for the prevention and control of the COVID-19 transmission. A novel molecular diagnosis technique, named multiplex reverse transcription loop-mediated isothermal amplification, that has been linked to a nanoparticle-based lateral flow biosensor (mRT-LAMP-LFB) was applied to detect SARS-CoV-2 based on the SARS-CoV-2 RdRp and N genes, and the mRT-LAMP products were analyzed using nanoparticle-based lateral flow biosensor. The mRT-LAMP-LFB amplification conditions, including the target RNA concentration, amplification temperature, and time were optimized. The sensitivity and specificity of the mRT-LAMP-LFB method were tested in the current study, and the mRT-LAMP-LFB assay was applied to detect the SARS-CoV-2 virus from clinical samples and artificial sputum samples. The SARS-CoV-2 specific primers based on the RdRp and N genes were valid for the establishment of mRT-LAMP-LFB assay to detect the SARS-CoV-2 virus. The multiple-RT-LAMP amplification condition was optimized at 63°C for 30 min. The full process, including reaction preparation, viral RNA extraction, RT-LAMP, and product identification, could be achieved in 80 min. The limit of detection (LoD) of the mRT-LAMP-LFB technology was 20 copies per reaction. The specificity of mRT-LAMP-LFB detection was 100%, and no cross-reactions to other respiratory pathogens were observed. The mRT-LAMP-LFB technique developed in the current study is a simple, rapid, and reliable method with great specificity and sensitivity when it comes to identifying SARS-CoV-2 virus for prevention and control of the COVID-19 disease, especially in resource-constrained regions of the world.
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