CRISPR/Cas9-Based Lateral Flow and Fluorescence Diagnostics

Mark J. Osborn; Akshay Bhardwaj; Samuel P. Bingea; Friederike Knipping; Colby J. Feser; Christopher J. Lees; Daniel P. Collins; Clifford J. Steer; Bruce R. Blazar; Jakub Tolar

doi:10.3390/bioengineering8020023

Bioengineering (Feb 2021)

CRISPR/Cas9-Based Lateral Flow and Fluorescence Diagnostics

Mark J. Osborn,
Akshay Bhardwaj,
Samuel P. Bingea,
Friederike Knipping,
Colby J. Feser,
Christopher J. Lees,
Daniel P. Collins,
Clifford J. Steer,
Bruce R. Blazar,
Jakub Tolar

Affiliations

Mark J. Osborn: Department of Pediatrics, Division of Blood and Marrow Transplant & Cellular Therapy, University of Minnesota Medical School, Minneapolis, MN 55455, USA
Akshay Bhardwaj: Department of Pediatrics, Division of Blood and Marrow Transplant & Cellular Therapy, University of Minnesota Medical School, Minneapolis, MN 55455, USA
Samuel P. Bingea: Department of Pediatrics, Division of Blood and Marrow Transplant & Cellular Therapy, University of Minnesota Medical School, Minneapolis, MN 55455, USA
Friederike Knipping: Department of Pediatrics, Division of Blood and Marrow Transplant & Cellular Therapy, University of Minnesota Medical School, Minneapolis, MN 55455, USA
Colby J. Feser: Department of Pediatrics, Division of Blood and Marrow Transplant & Cellular Therapy, University of Minnesota Medical School, Minneapolis, MN 55455, USA
Christopher J. Lees: Department of Pediatrics, Division of Blood and Marrow Transplant & Cellular Therapy, University of Minnesota Medical School, Minneapolis, MN 55455, USA
Daniel P. Collins: Cytomedical Design Group, LLC, Saint Paul, MN 55127, USA
Clifford J. Steer: Department of Medicine, University of Minnesota Medical School, Minneapolis, MN 55455, USA
Bruce R. Blazar: Department of Pediatrics, Division of Blood and Marrow Transplant & Cellular Therapy, University of Minnesota Medical School, Minneapolis, MN 55455, USA
Jakub Tolar: Department of Pediatrics, Division of Blood and Marrow Transplant & Cellular Therapy, University of Minnesota Medical School, Minneapolis, MN 55455, USA

DOI: https://doi.org/10.3390/bioengineering8020023
Journal volume & issue: Vol. 8, no. 2
p. 23

Abstract

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Clustered regularly interspaced short palindromic repeat (CRISPR/Cas) proteins can be designed to bind specified DNA and RNA sequences and hold great promise for the accurate detection of nucleic acids for diagnostics. We integrated commercially available reagents into a CRISPR/Cas9-based lateral flow assay that can detect severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) sequences with single-base specificity. This approach requires minimal equipment and represents a simplified platform for field-based deployment. We also developed a rapid, multiplex fluorescence CRISPR/Cas9 nuclease cleavage assay capable of detecting and differentiating SARS-CoV-2, influenza A and B, and respiratory syncytial virus in a single reaction. Our findings provide proof-of-principle for CRISPR/Cas9 point-of-care diagnosis as well as a scalable fluorescent platform for identifying respiratory viral pathogens with overlapping symptomology.

Published in Bioengineering

ISSN: 2306-5354 (Online)
Publisher: MDPI AG
Country of publisher: Switzerland
LCC subjects: Technology; Science: Biology (General)
Website: https://www.mdpi.com/journal/bioengineering

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Abstract

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