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A Simple, Affordable, Rapid, Stabilized, Colorimetric, Versatile RT-LAMP Assay to Detect SARS-CoV-2

The SARS-CoV-2 pandemic has forced all countries worldwide to rapidly develop and implement widespread testing to control and manage the Coronavirus Disease 2019 (COVID-19). reverse-transcription (RT)-qPCR is the gold standard molecular diagnostic method for COVID-19, mostly in automated testing pla...

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Autores principales: García-Bernalt Diego, Juan, Fernández-Soto, Pedro, Domínguez-Gil, Marta, Belhassen-García, Moncef, Bellido, Juan Luis Muñoz, Muro, Antonio
Formato: Online Artículo Texto
Lenguaje:English
Publicado: MDPI 2021
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8000859/
https://www.ncbi.nlm.nih.gov/pubmed/33806456
http://dx.doi.org/10.3390/diagnostics11030438
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author García-Bernalt Diego, Juan
Fernández-Soto, Pedro
Domínguez-Gil, Marta
Belhassen-García, Moncef
Bellido, Juan Luis Muñoz
Muro, Antonio
author_facet García-Bernalt Diego, Juan
Fernández-Soto, Pedro
Domínguez-Gil, Marta
Belhassen-García, Moncef
Bellido, Juan Luis Muñoz
Muro, Antonio
author_sort García-Bernalt Diego, Juan
collection PubMed
description The SARS-CoV-2 pandemic has forced all countries worldwide to rapidly develop and implement widespread testing to control and manage the Coronavirus Disease 2019 (COVID-19). reverse-transcription (RT)-qPCR is the gold standard molecular diagnostic method for COVID-19, mostly in automated testing platforms. These systems are accurate and effective, but also costly, time-consuming, high-technological, infrastructure-dependent, and currently suffer from commercial reagent supply shortages. The reverse-transcription loop-mediated isothermal amplification (RT-LAMP) can be used as an alternative testing method. Here, we present a novel versatile (real-time and colorimetric) RT-LAMP for the simple (one-step), affordable (~1.7 €/sample), and rapid detection of SARS-CoV-2 targeting both ORF1ab and N genes of the novel virus genome. We demonstrate the assay on RT-qPCR-positive clinical samples, obtaining most positive results under 25 min. In addition, a novel 30-min one-step drying protocol has been developed to stabilize the RT-LAMP reaction mixtures, allowing them to be stored at room temperature functionally for up to two months, as predicted by the Q(10). This Dry-RT-LAMP methodology is suitable for potentially ready-to-use COVID-19 diagnosis. After further testing and validation, it could be easily applied both in developed and in low-income countries yielding rapid and reliable results.
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spelling pubmed-80008592021-03-28 A Simple, Affordable, Rapid, Stabilized, Colorimetric, Versatile RT-LAMP Assay to Detect SARS-CoV-2 García-Bernalt Diego, Juan Fernández-Soto, Pedro Domínguez-Gil, Marta Belhassen-García, Moncef Bellido, Juan Luis Muñoz Muro, Antonio Diagnostics (Basel) Article The SARS-CoV-2 pandemic has forced all countries worldwide to rapidly develop and implement widespread testing to control and manage the Coronavirus Disease 2019 (COVID-19). reverse-transcription (RT)-qPCR is the gold standard molecular diagnostic method for COVID-19, mostly in automated testing platforms. These systems are accurate and effective, but also costly, time-consuming, high-technological, infrastructure-dependent, and currently suffer from commercial reagent supply shortages. The reverse-transcription loop-mediated isothermal amplification (RT-LAMP) can be used as an alternative testing method. Here, we present a novel versatile (real-time and colorimetric) RT-LAMP for the simple (one-step), affordable (~1.7 €/sample), and rapid detection of SARS-CoV-2 targeting both ORF1ab and N genes of the novel virus genome. We demonstrate the assay on RT-qPCR-positive clinical samples, obtaining most positive results under 25 min. In addition, a novel 30-min one-step drying protocol has been developed to stabilize the RT-LAMP reaction mixtures, allowing them to be stored at room temperature functionally for up to two months, as predicted by the Q(10). This Dry-RT-LAMP methodology is suitable for potentially ready-to-use COVID-19 diagnosis. After further testing and validation, it could be easily applied both in developed and in low-income countries yielding rapid and reliable results. MDPI 2021-03-04 /pmc/articles/PMC8000859/ /pubmed/33806456 http://dx.doi.org/10.3390/diagnostics11030438 Text en © 2021 by the authors. https://creativecommons.org/licenses/by/4.0/Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (http://creativecommons.org/licenses/by/4.0/ (https://creativecommons.org/licenses/by/4.0/) ).
spellingShingle Article
García-Bernalt Diego, Juan
Fernández-Soto, Pedro
Domínguez-Gil, Marta
Belhassen-García, Moncef
Bellido, Juan Luis Muñoz
Muro, Antonio
A Simple, Affordable, Rapid, Stabilized, Colorimetric, Versatile RT-LAMP Assay to Detect SARS-CoV-2
title A Simple, Affordable, Rapid, Stabilized, Colorimetric, Versatile RT-LAMP Assay to Detect SARS-CoV-2
title_full A Simple, Affordable, Rapid, Stabilized, Colorimetric, Versatile RT-LAMP Assay to Detect SARS-CoV-2
title_fullStr A Simple, Affordable, Rapid, Stabilized, Colorimetric, Versatile RT-LAMP Assay to Detect SARS-CoV-2
title_full_unstemmed A Simple, Affordable, Rapid, Stabilized, Colorimetric, Versatile RT-LAMP Assay to Detect SARS-CoV-2
title_short A Simple, Affordable, Rapid, Stabilized, Colorimetric, Versatile RT-LAMP Assay to Detect SARS-CoV-2
title_sort simple, affordable, rapid, stabilized, colorimetric, versatile rt-lamp assay to detect sars-cov-2
topic Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8000859/
https://www.ncbi.nlm.nih.gov/pubmed/33806456
http://dx.doi.org/10.3390/diagnostics11030438
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