A semi-automated, isolation-free, high-throughput SARS-CoV-2 reverse transcriptase (RT) loop-mediated isothermal amplification (LAMP) test

Abstract Shortages of reverse transcriptase (RT)-polymerase chain reaction (PCR) reagents and related equipment during the COVID-19 pandemic have demonstrated the need for alternative, high-throughput methods for severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2)-mass screening in clinical...

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Autores principales:	Jonas Schmidt, Sandro Berghaus, Frithjof Blessing, Folker Wenzel, Holger Herbeck, Josef Blessing, Peter Schierack, Stefan Rödiger, Dirk Roggenbuck
Formato:	article
Lenguaje:	EN
Publicado:	Nature Portfolio 2021
Materias:	Medicine R Science Q
Acceso en línea:	https://doaj.org/article/1fc9aa26de3f4534b3004dc1e4114f72
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spelling	oai:doaj.org-article:1fc9aa26de3f4534b3004dc1e4114f722021-11-08T10:50:55ZA semi-automated, isolation-free, high-throughput SARS-CoV-2 reverse transcriptase (RT) loop-mediated isothermal amplification (LAMP) test10.1038/s41598-021-00827-02045-2322https://doaj.org/article/1fc9aa26de3f4534b3004dc1e4114f722021-11-01T00:00:00Zhttps://doi.org/10.1038/s41598-021-00827-0https://doaj.org/toc/2045-2322Abstract Shortages of reverse transcriptase (RT)-polymerase chain reaction (PCR) reagents and related equipment during the COVID-19 pandemic have demonstrated the need for alternative, high-throughput methods for severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2)-mass screening in clinical diagnostic laboratories. A robust, SARS-CoV-2 RT-loop-mediated isothermal amplification (RT-LAMP) assay with high-throughput and short turnaround times in a clinical laboratory setting was established and compared to two conventional RT-PCR protocols using 323 samples of individuals with suspected SARS-CoV-2 infection. Limit of detection (LoD) and reproducibility of the isolation-free SARS-CoV-2 RT-LAMP test were determined. An almost perfect agreement (Cohen’s kappa > 0.8) between the novel test and two classical RT-PCR protocols with no systematic difference (McNemar’s test, P > 0.05) was observed. Sensitivity and specificity were in the range of 89.5 to 100% and 96.2 to 100% dependent on the reaction condition and the RT-PCR method used as reference. The isolation-free RT-LAMP assay showed high reproducibility (Tt intra-run coefficient of variation [CV] = 0.4%, Tt inter-run CV = 2.1%) with a LoD of 95 SARS-CoV-2 genome copies per reaction. The established SARS-CoV-2 RT-LAMP assay is a flexible and efficient alternative to conventional RT-PCR protocols, suitable for SARS-CoV-2 mass screening using existing laboratory infrastructure in clinical diagnostic laboratories.Jonas SchmidtSandro BerghausFrithjof BlessingFolker WenzelHolger HerbeckJosef BlessingPeter SchierackStefan RödigerDirk RoggenbuckNature PortfolioarticleMedicineRScienceQENScientific Reports, Vol 11, Iss 1, Pp 1-9 (2021)
institution	DOAJ
collection	DOAJ
language	EN
topic	Medicine R Science Q
spellingShingle	Medicine R Science Q Jonas Schmidt Sandro Berghaus Frithjof Blessing Folker Wenzel Holger Herbeck Josef Blessing Peter Schierack Stefan Rödiger Dirk Roggenbuck A semi-automated, isolation-free, high-throughput SARS-CoV-2 reverse transcriptase (RT) loop-mediated isothermal amplification (LAMP) test
description	Abstract Shortages of reverse transcriptase (RT)-polymerase chain reaction (PCR) reagents and related equipment during the COVID-19 pandemic have demonstrated the need for alternative, high-throughput methods for severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2)-mass screening in clinical diagnostic laboratories. A robust, SARS-CoV-2 RT-loop-mediated isothermal amplification (RT-LAMP) assay with high-throughput and short turnaround times in a clinical laboratory setting was established and compared to two conventional RT-PCR protocols using 323 samples of individuals with suspected SARS-CoV-2 infection. Limit of detection (LoD) and reproducibility of the isolation-free SARS-CoV-2 RT-LAMP test were determined. An almost perfect agreement (Cohen’s kappa > 0.8) between the novel test and two classical RT-PCR protocols with no systematic difference (McNemar’s test, P > 0.05) was observed. Sensitivity and specificity were in the range of 89.5 to 100% and 96.2 to 100% dependent on the reaction condition and the RT-PCR method used as reference. The isolation-free RT-LAMP assay showed high reproducibility (Tt intra-run coefficient of variation [CV] = 0.4%, Tt inter-run CV = 2.1%) with a LoD of 95 SARS-CoV-2 genome copies per reaction. The established SARS-CoV-2 RT-LAMP assay is a flexible and efficient alternative to conventional RT-PCR protocols, suitable for SARS-CoV-2 mass screening using existing laboratory infrastructure in clinical diagnostic laboratories.
format	article
author	Jonas Schmidt Sandro Berghaus Frithjof Blessing Folker Wenzel Holger Herbeck Josef Blessing Peter Schierack Stefan Rödiger Dirk Roggenbuck
author_facet	Jonas Schmidt Sandro Berghaus Frithjof Blessing Folker Wenzel Holger Herbeck Josef Blessing Peter Schierack Stefan Rödiger Dirk Roggenbuck
author_sort	Jonas Schmidt
title	A semi-automated, isolation-free, high-throughput SARS-CoV-2 reverse transcriptase (RT) loop-mediated isothermal amplification (LAMP) test
title_short	A semi-automated, isolation-free, high-throughput SARS-CoV-2 reverse transcriptase (RT) loop-mediated isothermal amplification (LAMP) test
title_full	A semi-automated, isolation-free, high-throughput SARS-CoV-2 reverse transcriptase (RT) loop-mediated isothermal amplification (LAMP) test
title_fullStr	A semi-automated, isolation-free, high-throughput SARS-CoV-2 reverse transcriptase (RT) loop-mediated isothermal amplification (LAMP) test
title_full_unstemmed	A semi-automated, isolation-free, high-throughput SARS-CoV-2 reverse transcriptase (RT) loop-mediated isothermal amplification (LAMP) test
title_sort	semi-automated, isolation-free, high-throughput sars-cov-2 reverse transcriptase (rt) loop-mediated isothermal amplification (lamp) test
publisher	Nature Portfolio
publishDate	2021
url	https://doaj.org/article/1fc9aa26de3f4534b3004dc1e4114f72
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A semi-automated, isolation-free, high-throughput SARS-CoV-2 reverse transcriptase (RT) loop-mediated isothermal amplification (LAMP) test

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