Title: Amplification-free detection of SARS-CoV-2 with CRISPR-Cas13a and mobile phone microscopy
Author: Parinaz Fozouni, Sungmin Son, María Díaz de León Derby, Gavin J. Knott, Carley N. Gray, Michael V. D’Ambrosio, Chunyu Zhao, Neil A. Switz, G. Renuka Kumar, Stephanie I. Stephens, Daniela Boehm, Chia-Lin Tsou, Jeffrey Shu, Abdul Bhuiya, Max Armstrong, Andrew R. Harris, Pei-Yi Chen, Jeannette M. Osterloh, Anke Meyer-Franke, Bastian Joehnk, Keith Walcott, Anita Sil, Charles Langelier, Katherine S. Pollard, Emily D. Crawford, Andreas S. Puschnik, Maira Phelps, Amy Kistler, Joseph L. DeRisi, Jennifer A. Doudna, Daniel A. Fletcher, Melanie Ott
Abstract: The December 2019 outbreak of a novel respiratory virus, SARS-CoV-2, has become an ongoing global pandemic due in part to the challenge of identifying symptomatic, asymptomatic and pre-symptomatic carriers of the virus. CRISPR diagnostics can augment gold-standard PCR-based testing if they can be made rapid, portable and accurate. Here we report the development of an amplification-free CRISPR-Cas13a assay for direct detection of SARS-CoV-2 from nasal swab RNA that can be read with a mobile phone microscope. The assay achieved ～100 copies/μL sensitivity in under 30 minutes of measurement time and accurately detected pre-extracted RNA from a set of positive clinical samples in under 5 minutes. We combined crRNAs targeting SARS-CoV-2 RNA to improve sensitivity and specificity, and directly quantified viral load using enzyme kinetics. Integrated with a reader device based on a mobile phone, this assay has the potential to enable rapid, low-cost, point-of-care screening for SARS-CoV-2.