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Rapid SARS-CoV-2 testing in primary material based on a novel multiplex RT-LAMP assay.
Schermer, Bernhard; Fabretti, Francesca; Damagnez, Maximilian; Di Cristanziano, Veronica; Heger, Eva; Arjune, Sita; Tanner, Nathan A; Imhof, Thomas; Koch, Manuel; Ladha, Alim; Joung, Julia; Gootenberg, Jonathan S; Abudayyeh, Omar O; Burst, Volker; Zhang, Feng; Klein, Florian; Benzing, Thomas; Müller, Roman-Ulrich.
  • Schermer B; Department II of Internal Medicine, University of Cologne, Faculty of Medicine and University Hospital Cologne, Cologne, Germany.
  • Fabretti F; Center for Molecular Medicine Cologne, University of Cologne, Faculty of Medicine and University Hospital Cologne, Cologne, Germany.
  • Damagnez M; Cologne Excellence Cluster on Cellular Stress Responses in Aging-Associated Diseases (CECAD), University of Cologne, Cologne, Germany.
  • Di Cristanziano V; Department II of Internal Medicine, University of Cologne, Faculty of Medicine and University Hospital Cologne, Cologne, Germany.
  • Heger E; Center for Molecular Medicine Cologne, University of Cologne, Faculty of Medicine and University Hospital Cologne, Cologne, Germany.
  • Arjune S; Cologne Excellence Cluster on Cellular Stress Responses in Aging-Associated Diseases (CECAD), University of Cologne, Cologne, Germany.
  • Tanner NA; Institute of Virology, Faculty of Medicine and University Hospital Cologne, University of Cologne, Cologne, Germany.
  • Imhof T; Institute of Virology, Faculty of Medicine and University Hospital Cologne, University of Cologne, Cologne, Germany.
  • Koch M; Institute of Virology, Faculty of Medicine and University Hospital Cologne, University of Cologne, Cologne, Germany.
  • Ladha A; Department II of Internal Medicine, University of Cologne, Faculty of Medicine and University Hospital Cologne, Cologne, Germany.
  • Joung J; Center for Molecular Medicine Cologne, University of Cologne, Faculty of Medicine and University Hospital Cologne, Cologne, Germany.
  • Gootenberg JS; Cologne Excellence Cluster on Cellular Stress Responses in Aging-Associated Diseases (CECAD), University of Cologne, Cologne, Germany.
  • Abudayyeh OO; New England Biolabs, Ipswich, MA, United States of America.
  • Burst V; Center for Molecular Medicine Cologne, University of Cologne, Faculty of Medicine and University Hospital Cologne, Cologne, Germany.
  • Zhang F; Institute for Dental Research and Oral Musculoskeletal Biology, Center for Biochemistry, University of Cologne, Cologne, Germany.
  • Klein F; Center for Molecular Medicine Cologne, University of Cologne, Faculty of Medicine and University Hospital Cologne, Cologne, Germany.
  • Benzing T; Institute for Dental Research and Oral Musculoskeletal Biology, Center for Biochemistry, University of Cologne, Cologne, Germany.
  • Müller RU; Howard Hughes Medical Institute, Cambridge, MA, United States of America.
PLoS One ; 15(11): e0238612, 2020.
Article in English | MEDLINE | ID: covidwho-902011
Preprint
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ABSTRACT

BACKGROUND:

Rapid and extensive testing of large parts of the population and specific subgroups is crucial for proper management of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infections and decision-making in times of a pandemic outbreak. However, point-of-care (POC) testing in places such as emergency units, outpatient clinics, airport security points or the entrance of any public building is a major challenge. The need for thermal cycling and nucleic acid isolation hampers the use of standard PCR-based methods for this purpose.

METHODS:

To avoid these obstacles, we tested PCR-independent methods for the detection of SARS-CoV-2 RNA from primary material (nasopharyngeal swabs) including reverse transcription loop-mediated isothermal amplification (RT-LAMP) and specific high-sensitivity enzymatic reporter unlocking (SHERLOCK).

RESULTS:

Whilst specificity of standard RT-LAMP assays appears to be satisfactory, sensitivity does not reach the current gold-standard quantitative real-time polymerase chain reaction (qPCR) assays yet. We describe a novel multiplexed RT-LAMP approach and validate its sensitivity on primary samples. This approach allows for fast and reliable identification of infected individuals. Primer optimization and multiplexing helps to increase sensitivity significantly. In addition, we directly compare and combine our novel RT-LAMP assays with SHERLOCK.

CONCLUSION:

In summary, this approach reveals one-step multiplexed RT-LAMP assays as a prime-option for the development of easy and cheap POC test kits.
Subject(s)

Full text: Available Collection: International databases Database: MEDLINE Main subject: Pneumonia, Viral / RNA, Viral / Coronavirus Infections / Nucleic Acid Amplification Techniques / Betacoronavirus Type of study: Diagnostic study / Prognostic study Limits: Humans Language: English Journal: PLoS One Journal subject: Science / Medicine Year: 2020 Document Type: Article Affiliation country: Journal.pone.0238612

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Full text: Available Collection: International databases Database: MEDLINE Main subject: Pneumonia, Viral / RNA, Viral / Coronavirus Infections / Nucleic Acid Amplification Techniques / Betacoronavirus Type of study: Diagnostic study / Prognostic study Limits: Humans Language: English Journal: PLoS One Journal subject: Science / Medicine Year: 2020 Document Type: Article Affiliation country: Journal.pone.0238612