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Highly performing point-of-care molecular testing for SARS-CoV-2 with RNA extraction and isothermal amplification.

Résumé : In order to respond to the urgent request of massive testing, developed countries perform nucleic acid amplification tests (NAAT) of SARS-CoV-2 in centralized laboratories. Real-time RT - PCR (Reverse transcription - Polymerase Chain Reaction) is used to amplify the viral RNA and enable its detection. Although PCR is 37 years old, it is still considered, without dispute, as the gold standard. PCR is an efficient process, but the complex engineering required for automated RNA extraction and temperature cycling makes it incompatible for use in point of care settings. In the present work, by harnessing progress made in the past two decades in DNA amplification, microfluidics and membrane technologies, we succeeded to create a portable test, in which SARS-CoV-2 RNA is extracted, amplified isothermally by RT - LAMP (Loop-mediated Isothermal Amplification), and detected using intercalating dyes or highly fluorescent probes. Depending on the viral load, the detection takes between twenty minutes and one hour. Using pools of naso-pharyngal clinical samples, we estimated a sensitivity comparable to RT-qPCR (up to a Cycle threshold of 39, equivalent to <0.1 TCID50 per mL) and a 100% specificity, for other human coronaviruses and eight respiratory viruses currently circulating in Europe. We designed and fabricated an easy-to-use portable device called COVIDISC to carry out the test at the point of care. The low cost of the materials along with the absence of complex equipment paves the way towards a large dissemination of this device. The perspective of a reliable SARS-CoV-2 point of care detection, highly performing, that would deliver on-site results in less than one hour opens up a new efficient approach to manage the pandemics.
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https://hal-pasteur.archives-ouvertes.fr/pasteur-02916049
Contributor : Jessica Vanhomwegen <>
Submitted on : Friday, August 21, 2020 - 1:20:28 PM
Last modification on : Saturday, September 26, 2020 - 11:44:11 PM

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Pierre Garneret, Etienne Coz, Elian Martin, Jean-Claude Manuguerra, Elodie Brient-Litzler, et al.. Highly performing point-of-care molecular testing for SARS-CoV-2 with RNA extraction and isothermal amplification.. 2020. ⟨pasteur-02916049⟩

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