Maturada PatchsungKrittapas JantarugArchiraya PattamaKanokpol AphichoSurased SuraritdechachaiPiyachat MeesawatKhomkrit SappakhawNattawat LeelahakornTheerawat RuenkamThanakrit WongsatitNiracha AthipanyasilpBhumrapee EiamthongBenya LakkanasiroratThitima PhoodokmaiNootaree NiljianskulDanaya PakotipraphaSittinan ChanaratAimorn HomchanRuchanok TinikulPhilaiwarong KamutiraKochakorn PhiwkaowSahachat SoithongcharoenChadaporn KantiwiriyawanitchVinutsada PongsupasaDuangthip TrisriviratJuthamas JaroensukThanyaporn WongnateSomchart MaenpuenPimchai ChaiyenSirichai KamnerdnaktaJirawat SwangsriSuebwong ChuthapisithYongyut SirivatanauksornChutikarn ChaimayoRuengpung SutthentWannee KantakamalakulJulia JoungAlim LadhaXin JinJonathan S. GootenbergOmar O. AbudayyehFeng ZhangNavin HorthongkhamChayasith UttamapinantPTT Public Company LimitedVidyasirimedhi Institute of Science and TechnologyMcGovern InstituteSociety of Fellows, Harvard UniversityMassachusetts Institute of TechnologyMahidol UniversityFaculty of Medicine, Siriraj Hospital, Mahidol UniversityHarvard UniversityHoward Hughes Medical InstituteBurapha UniversityBroad InstituteMassachusetts Consortium on Pathogen Readiness2020-10-052020-10-052020-01-01Nature Biomedical Engineering. (2020)2157846X2-s2.0-85089869548https://repository.li.mahidol.ac.th/handle/20.500.14594/59008© 2020, The Author(s), under exclusive licence to Springer Nature Limited. Nucleic acid detection by isothermal amplification and the collateral cleavage of reporter molecules by CRISPR-associated enzymes is a promising alternative to quantitative PCR. Here, we report the clinical validation of the specific high-sensitivity enzymatic reporter unlocking (SHERLOCK) assay using the enzyme Cas13a from Leptotrichia wadei for the detection of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2)—the virus that causes coronavirus disease 2019 (COVID-19)—in 154 nasopharyngeal and throat swab samples collected at Siriraj Hospital, Thailand. Within a detection limit of 42 RNA copies per reaction, SHERLOCK was 100% specific and 100% sensitive with a fluorescence readout, and 100% specific and 97% sensitive with a lateral-flow readout. For the full range of viral load in the clinical samples, the fluorescence readout was 100% specific and 96% sensitive. For 380 SARS-CoV-2-negative pre-operative samples from patients undergoing surgery, SHERLOCK was in 100% agreement with quantitative PCR with reverse transcription. The assay, which we show is amenable to multiplexed detection in a single lateral-flow strip incorporating an internal control for ribonuclease contamination, should facilitate SARS-CoV-2 detection in settings with limited resources.Mahidol UniversityBiochemistry, Genetics and Molecular BiologyChemical EngineeringComputer ScienceEngineeringMedicineArticleSCOPUS10.1038/s41551-020-00603-x