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Scalable, rapid and highly sensitive isothermal detection of SARS-CoV-2 for laboratory and home testing

Max J. Kellner, James J. Ross, Jakob Schnabl, Marcus P.S. Dekens, Robert Heinen, Nathan A. Tanner, Robert Fritsche-Polanz, Marianna Traugott, Tamara Seitz, Alexander Zoufaly, Manuela Födinger, Christoph Wenisch, Johannes Zuber, Vienna Covid-19 Diagnostics Initiative (VCDI), View ORCID ProfileAndrea Pauli, Julius Brennecke
doi: https://doi.org/10.1101/2020.06.23.166397
Max J. Kellner
1Research Institute of Molecular Pathology (IMP), Vienna BioCenter (VBC), Campus-Vienna-Biocenter 1, 1030 Vienna, Austria
2Institute of Molecular Biotechnology of the Austrian Academy of Sciences (IMBA), Vienna BioCenter (VBC), Dr. Bohr-Gasse 3, 1030 Vienna, Austria
3MRC Laboratory of Molecular Biology, Francis Crick Avenue, Cambridge CB2 0QH, UK
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  • For correspondence: mkellner@mrc-lmb.cam.ac.uk andrea.pauli@imp.ac.at julius.brennecke@imba.oeaw.ac.at
James J. Ross
2Institute of Molecular Biotechnology of the Austrian Academy of Sciences (IMBA), Vienna BioCenter (VBC), Dr. Bohr-Gasse 3, 1030 Vienna, Austria
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Jakob Schnabl
2Institute of Molecular Biotechnology of the Austrian Academy of Sciences (IMBA), Vienna BioCenter (VBC), Dr. Bohr-Gasse 3, 1030 Vienna, Austria
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Marcus P.S. Dekens
1Research Institute of Molecular Pathology (IMP), Vienna BioCenter (VBC), Campus-Vienna-Biocenter 1, 1030 Vienna, Austria
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Robert Heinen
1Research Institute of Molecular Pathology (IMP), Vienna BioCenter (VBC), Campus-Vienna-Biocenter 1, 1030 Vienna, Austria
2Institute of Molecular Biotechnology of the Austrian Academy of Sciences (IMBA), Vienna BioCenter (VBC), Dr. Bohr-Gasse 3, 1030 Vienna, Austria
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Nathan A. Tanner
4New England Biolabs (NEB), 240 County Road, Ipswich, MA, 01938, USA
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Robert Fritsche-Polanz
5Institute of Laboratory Diagnostics, Klinik Favoriten, 1100 Vienna, Austria
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Marianna Traugott
6th, 1100 Vienna, Austria
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Tamara Seitz
6th, 1100 Vienna, Austria
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Alexander Zoufaly
6th, 1100 Vienna, Austria
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Manuela Födinger
5Institute of Laboratory Diagnostics, Klinik Favoriten, 1100 Vienna, Austria
7Sigmund Freud Private University, 1020 Vienna, Austria
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Christoph Wenisch
6th, 1100 Vienna, Austria
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Johannes Zuber
1Research Institute of Molecular Pathology (IMP), Vienna BioCenter (VBC), Campus-Vienna-Biocenter 1, 1030 Vienna, Austria
8Medical University of Vienna, Vienna BioCenter (VBC), 1030 Vienna, Austria
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Andrea Pauli
1Research Institute of Molecular Pathology (IMP), Vienna BioCenter (VBC), Campus-Vienna-Biocenter 1, 1030 Vienna, Austria
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  • ORCID record for Andrea Pauli
  • For correspondence: mkellner@mrc-lmb.cam.ac.uk andrea.pauli@imp.ac.at julius.brennecke@imba.oeaw.ac.at
Julius Brennecke
2Institute of Molecular Biotechnology of the Austrian Academy of Sciences (IMBA), Vienna BioCenter (VBC), Dr. Bohr-Gasse 3, 1030 Vienna, Austria
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  • For correspondence: mkellner@mrc-lmb.cam.ac.uk andrea.pauli@imp.ac.at julius.brennecke@imba.oeaw.ac.at
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Abstract

Global efforts to monitor and contain the Covid-19 pandemic, caused by the beta-coronavirus SARS-CoV-2, currently rely on RT-qPCR-based diagnostic assays. Yet their high cost, moderate throughput, and dependence on sophisticated equipment limit a broad implementation. Loop-mediated isothermal amplification (RT-LAMP) is an alternative detection method that has the potential to overcome these limitations. Here, we established a robust, highly sensitive and versatile RT-LAMP-based SARS-CoV-2 detection assay that is insensitive to carry-over contaminations. Our approach uses a rapid upfront lysis step and hydroxy-naphthol-blue (HNB) for colorimetric detection, which enables the robust identification of Covid-19 infections from a variety of sample types within 30 minutes. By combining RT-LAMP with a simple nucleic acid enrichment method (bead-LAMP), we profoundly increased assay sensitivity to RT-qPCR-like levels, thereby extending applications to large-scale pooled testing. Finally, we developed HomeDip-LAMP for pipette-free SARS-CoV-2 detection for low-resource environments. Our combined optimizations set the stage for implementing RT-LAMP as SARS-CoV-2 diagnostics assay for population-wide and home-based testing.

Competing Interest Statement

NAT is employee and stockholder of New England Biolabs, the manufacturer of reagents used in the study. All other authors declare no conflict of interest.

Copyright 
The copyright holder for this preprint is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is made available under a CC-BY-NC-ND 4.0 International license.
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Posted June 23, 2020.
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Scalable, rapid and highly sensitive isothermal detection of SARS-CoV-2 for laboratory and home testing
Max J. Kellner, James J. Ross, Jakob Schnabl, Marcus P.S. Dekens, Robert Heinen, Nathan A. Tanner, Robert Fritsche-Polanz, Marianna Traugott, Tamara Seitz, Alexander Zoufaly, Manuela Födinger, Christoph Wenisch, Johannes Zuber, Vienna Covid-19 Diagnostics Initiative (VCDI), Andrea Pauli, Julius Brennecke
bioRxiv 2020.06.23.166397; doi: https://doi.org/10.1101/2020.06.23.166397
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Scalable, rapid and highly sensitive isothermal detection of SARS-CoV-2 for laboratory and home testing
Max J. Kellner, James J. Ross, Jakob Schnabl, Marcus P.S. Dekens, Robert Heinen, Nathan A. Tanner, Robert Fritsche-Polanz, Marianna Traugott, Tamara Seitz, Alexander Zoufaly, Manuela Födinger, Christoph Wenisch, Johannes Zuber, Vienna Covid-19 Diagnostics Initiative (VCDI), Andrea Pauli, Julius Brennecke
bioRxiv 2020.06.23.166397; doi: https://doi.org/10.1101/2020.06.23.166397

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