Clinical validation of a Cas13-based assay for the detection of SARS-CoV-2 RNA

被引：493

作者：

Patchsung, Maturada ^{[1
]}

Jantarug, Krittapas ^{[1
]}

Pattama, Archiraya ^{[2
]}

Aphicho, Kanokpol ^{[1
]}

Suraritdechachai, Surased ^{[1
]}

Meesawat, Piyachat ^{[1
]}

Sappakhaw, Khomkrit ^{[1
]}

Leelahakorn, Nattawat ^{[1
]}

Ruenkam, Theerawat ^{[1
]}

Wongsatit, Thanakrit ^{[1
]}

Athipanyasilp, Niracha ^{[2
]}

Eiamthong, Bhumrapee ^{[1
]}

Lakkanasirorat, Benya ^{[1
]}

Phoodokmai, Thitima ^{[1
]}

Niljianskul, Nootaree ^{[3
]}

Pakotiprapha, Danaya ^{[4
,5
]}

Chanarat, Sittinan ^{[4
,5
]}

Homchan, Aimorn ^{[4
,5
]}

Tinikul, Ruchanok ^{[4
,5
]}

Kamutira, Philaiwarong ^{[4
,5
]}

Phiwkaow, Kochakorn ^{[1
]}

Soithongcharoen, Sahachat ^{[1
]}

Kantiwiriyawanitch, Chadaporn ^{[1
]}

Pongsupasa, Vinutsada ^{[1
]}

Trisrivirat, Duangthip ^{[1
]}

Jaroensuk, Juthamas ^{[1
]}

Wongnate, Thanyaporn ^{[1
]}

Maenpuen, Somchart ^{[6
]}

Chaiyen, Pimchai ^{[1
]}

Kamnerdnakta, Sirichai ^{[7
]}

Swangsri, Jirawat ^{[7
]}

Chuthapisith, Suebwong ^{[7
]}

Sirivatanauksorn, Yongyut ^{[7
]}

Chaimayo, Chutikarn ^{[2
]}

Sutthent, Ruengpung ^{[2
]}

Kantakamalakul, Wannee ^{[2
]}

Joung, Julia ^{[8
,9
,10
,11
]}

Ladha, Alim ^{[8
,9
,10
,11
]}

Jin, Xin ^{[9
,10
,12
,13
]}

Gootenberg, Jonathan S. ^{[10
,14
]}

Abudayyeh, Omar O. ^{[10
,14
]}

Zhang, Feng ^{[8
,9
,10
,11
,14
,15
]}

Horthongkham, Navin ^{[2
]}

Uttamapinant, Chayasith ^{[1
]}

机构：

[1] Vidyasirimedhi Inst Sci & Technol VISTEC, Sch Biomol Sci & Engn, Rayong, Thailand

[2] Mahidol Univ, Fac Med, Dept Microbiol, Siriraj Hosp, Bangkok, Thailand

[3] PTT Publ Co Ltd, Bangkok, Thailand

[4] Mahidol Univ, Fac Sci, Dept Biochem, Bangkok, Thailand

[5] Mahidol Univ, Fac Sci, Ctr Excellence Prot & Enzyme Technol, Bangkok, Thailand

[6] Burapha Univ, Fac Sci, Dept Biochem, Chon Buri, Thailand

[7] Mahidol Univ, Fac Med, Dept Surg, Siriraj Hosp, Bangkok, Thailand

[8] Howard Hughes Med Inst, Cambridge, MA USA

[9] Broad Inst MIT & Harvard, Cambridge, MA 02142 USA

[10] MIT, McGovern Inst Brain Res, 77 Massachusetts Ave, Cambridge, MA 02139 USA

[11] MIT, Dept Biol Engn, 77 Massachusetts Ave, Cambridge, MA 02139 USA

[12] Harvard Univ, Soc Fellows, Cambridge, MA 02138 USA

[13] Harvard Univ, Dept Stem Cell & Regenerat Biol, Cambridge, MA 02138 USA

[14] Massachusetts Consortium Pathogen Readiness, Boston, MA USA

[15] MIT, Dept Brain & Cognit Sci, E25-618, Cambridge, MA 02139 USA

来源：

NATURE BIOMEDICAL ENGINEERING | 2020年 / 4卷 / 12期

关键词：

NUCLEIC-ACID DETECTION; SEQUENCE; PLATFORM;

D O I：

10.1038/s41551-020-00603-x

中图分类号：

R318 [生物医学工程];

学科分类号：

0831 ;

摘要：

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 fromLeptotrichia wadeifor 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. The specific high-sensitivity enzymatic reporter unlocking (SHERLOCK) assay detected severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) RNA with high sensitivity and specificity in hundreds of nasopharyngeal and throat swab samples collected at Siriraj Hospital in Thailand.

引用

页码：1140 / 1149

页数：10

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