Resilient SARS-CoV-2 diagnostics workflows including viral heat inactivation.
Resilient SARS-CoV-2 diagnostics workflows including viral heat inactivation.
复制标题
DOI：
10.1371/journal.pone.0256813
复制
发表时间：
2021
期刊：
影响因子：
3.7
通讯作者：
Martinez-Nunez RT
中科院分区：
文献类型：
--
作者：
Lista MJ;Matos PM;Maguire TJA;Poulton K;Ortiz-Zapater E;Page R;Sertkaya H;Ortega-Prieto AM;Scourfield E;O'Byrne AM;Bouton C;Dickenson RE;Ficarelli M;Jimenez-Guardeño JM;Howard M;Betancor G;Galao RP;Pickering S;Signell AW;Wilson H;Cliff P;Kia Ik MT;Patel A;MacMahon E;Cunningham E;Doores K;Agromayor M;Martin-Serrano J;Perucha E;Mischo HE;Shankar-Hari M;Batra R;Edgeworth J;Zuckerman M;Malim MH;Neil S;Martinez-Nunez RT
全球范围内都需要进行SARS-CoV-2检测的试剂。一些实验室已经实施了无试剂盒方案，但许多其他实验室没有能力开发这些方案和/或进行人工处理。我们通过比较几种市售的RNA提取方法，为临床样本中的SARS-CoV-2核酸检测提供了多种工作流程：QIAamp Viral RNA Mini Kit（QIAgen），RNAdvance Blood/Viral（Beckman）和Mag-Bind Viral DNA/RNA 96 Kit（Omega Bio-tek）。我们还比较了一步RT-qPCR试剂：TaqMan Fast Virus 1-Step Master Mix（FastVirus，ThermoFisher Scientific）、qPCRBIO Probe 1-Step Go Lo-ROX（PCR Biosystems）和Luna® Universal Probe One-Step RT-qPCR Kit（Luna，NEB）。我们使用检测病毒N（EUA CDC）和RdRP的引物探针。RNA提取方法提供了类似的结果，贝克曼与我们的引物探针组合表现更好。Luna被证明是最敏感的，尽管总体上三种试剂没有显示出显着差异。N检测比RdRP更可靠，特别是在低病毒滴度的样品中。重要的是，我们证明了在70°C下热处理鼻咽拭子10或30分钟，或90°C下热处理鼻咽拭子10或30分钟（原始变体和B 1.1.7）灭活SARS-CoV-2，采用空斑试验，对临床样本中qPCR的灵敏度影响最小。这些发现使SARS-CoV-2检测在没有CL-3设施的环境中变得便携。总之，我们提供了几个可以在其他实验室轻松实施的测试管道，并在https://osf.io/uebvj/上免费提供我们所有的协议和SOP。
There is a worldwide need for reagents to perform SARS-CoV-2 detection. Some laboratories have implemented kit-free protocols, but many others do not have the capacity to develop these and/or perform manual processing. We provide multiple workflows for SARS-CoV-2 nucleic acid detection in clinical samples by comparing several commercially available RNA extraction methods: QIAamp Viral RNA Mini Kit (QIAgen), RNAdvance Blood/Viral (Beckman) and Mag-Bind Viral DNA/RNA 96 Kit (Omega Bio-tek). We also compared One-step RT-qPCR reagents: TaqMan Fast Virus 1-Step Master Mix (FastVirus, ThermoFisher Scientific), qPCRBIO Probe 1-Step Go Lo-ROX (PCR Biosystems) and Luna® Universal Probe One-Step RT-qPCR Kit (Luna, NEB). We used primer-probes that detect viral N (EUA CDC) and RdRP. RNA extraction methods provided similar results, with Beckman performing better with our primer-probe combinations. Luna proved most sensitive although overall the three reagents did not show significant differences. N detection was more reliable than that of RdRP, particularly in samples with low viral titres. Importantly, we demonstrated that heat treatment of nasopharyngeal swabs at 70°C for 10 or 30 min, or 90°C for 10 or 30 min (both original variant and B 1.1.7) inactivated SARS-CoV-2 employing plaque assays, and had minimal impact on the sensitivity of the qPCR in clinical samples. These findings make SARS-CoV-2 testing portable in settings that do not have CL-3 facilities. In summary, we provide several testing pipelines that can be easily implemented in other laboratories and have made all our protocols and SOPs freely available at https://osf.io/uebvj/.
登录
查看更多内容
SARS-COV-2血统的估计可传播和影响B.1.1.7在英国。
DOI：
10.1126/science.abg3055
发表时间：
2021-04-09
期刊：
Science (New York, N.Y.)
影响因子：
--
作者：
Davies NG;Abbott S;Barnard RC;Jarvis CI;Kucharski AJ;Munday JD;Pearson CAB;Russell TW;Tully DC;Washburne AD;Wenseleers T;Gimma A;Waites W;Wong KLM;van Zandvoort K;Silverman JD;CMMID COVID-19 Working Group;COVID-19 Genomics UK (COG-UK) Consortium;Diaz-Ordaz K;Keogh R;Eggo RM;Funk S;Jit M;Atkins KE;Edmunds WJ
通讯作者：
Edmunds WJ
影响因子：
9.8
作者：
Reijns MAM;Thompson L;Acosta JC;Black HA;Sanchez-Luque FJ;Diamond A;Parry DA;Daniels A;O'Shea M;Uggenti C;Sanchez MC;O'Callaghan A;McNab MLL;Adamowicz M;Friman ET;Hurd T;Jarman EJ;Chee FLM;Rainger JK;Walker M;Drake C;Longman D;Mordstein C;Warlow SJ;McKay S;Slater L;Ansari M;Tomlinson IPM;Moore D;Wilkinson N;Shepherd J;Templeton K;Johannessen I;Tait-Burkard C;Haas JG;Gilbert N;Adams IR;Jackson AP
通讯作者：
Jackson AP
影响因子：
9.3
作者：
Pan, Yang;Long, Luyao;Ren, Simei
通讯作者：
Ren, Simei
评估SARS-COV-2的失活程序。
DOI：
10.1099/jgv.0.001539
发表时间：
2021-03
期刊：
The Journal of general virology
影响因子：
--
作者：
Auerswald H;Yann S;Dul S;In S;Dussart P;Martin NJ;Karlsson EA;Garcia-Rivera JA
通讯作者：
Garcia-Rivera JA
DOI：
10.1016/j.jcv.2020.104423
发表时间：
2020-07-01
期刊：
Journal of clinical virology : the official publication of the Pan American Society for Clinical Virology
影响因子：
--
作者：
Merindol, Natacha;Pepin, Genevieve;Danylo, Alexis
通讯作者：
Danylo, Alexis