{"database":"biostudies-literature","file_versions":[],"scores":null,"additional":{"submitter":["Nguyen LT"],"funding":["CGH CDC HHS","NIAID NIH HHS"],"pagination":["7"],"full_dataset_link":["https://www.ebi.ac.uk/biostudies/studies/S-EPMC9053293"],"repository":["biostudies-literature"],"omics_type":["Unknown"],"volume":["2"],"pubmed_abstract":["<h4>Background</h4>The coronavirus disease (COVID-19) caused by SARS-CoV-2 has swept through the globe at an unprecedented rate. CRISPR-based detection technologies have emerged as a rapid and affordable platform that can shape the future of diagnostics.<h4>Methods</h4>We developed ENHANCEv2 that is composed of a chimeric guide RNA, a modified LbCas12a enzyme, and a dual reporter construct to improve the previously reported ENHANCE system. We validated both ENHANCE and ENHANCEv2 using 62 nasopharyngeal swabs and compared the results to RT-qPCR. We created a lyophilized version of ENHANCEv2 and characterized its detection capability and stability.<h4>Results</h4>Here we demonstrate that when coupled with an RT-LAMP step, ENHANCE detects COVID-19 samples down to a few copies with 95% accuracy while maintaining a high specificity towards various isolates of SARS-CoV-2 against 31 highly similar and common respiratory pathogens. ENHANCE works robustly in a wide range of magnesium concentrations (3 mM-13 mM), allowing for further assay optimization. Our clinical validation results for both ENHANCE and ENHANCEv2 show 60/62 (96.7%) sample agreement with RT-qPCR results while only using 5 µL of sample and 20 minutes of CRISPR reaction. We show that the lateral flow assay using paper-based strips displays 100% agreement with the fluorescence-based reporter assay during clinical validation. Finally, we demonstrate that a lyophilized version of ENHANCEv2 shows high sensitivity and specificity for SARS-CoV-2 detection while reducing the CRISPR reaction time to as low as 3 minutes while maintaining its detection capability for several weeks upon storage at room temperature.<h4>Conclusions</h4>CRISPR-based diagnostic platforms offer many advantages as compared to conventional qPCR-based detection methods. Our work here provides clinical validation of ENHANCE and its improved form ENHANCEv2 for the detection of COVID-19."],"journal":["Communications medicine"],"pubmed_title":["Clinical validation of engineered CRISPR/Cas12a for rapid SARS-CoV-2 detection."],"pmcid":["PMC9053293"],"funding_grant_id":["U01 GH002338","R21 AI156321"],"pubmed_authors":["Jain PK","Rananaware SR","Stone BT","Nguyen LT","Pizzano BLM"],"additional_accession":[]},"is_claimable":false,"name":"Clinical validation of engineered CRISPR/Cas12a for rapid SARS-CoV-2 detection.","description":"<h4>Background</h4>The coronavirus disease (COVID-19) caused by SARS-CoV-2 has swept through the globe at an unprecedented rate. CRISPR-based detection technologies have emerged as a rapid and affordable platform that can shape the future of diagnostics.<h4>Methods</h4>We developed ENHANCEv2 that is composed of a chimeric guide RNA, a modified LbCas12a enzyme, and a dual reporter construct to improve the previously reported ENHANCE system. We validated both ENHANCE and ENHANCEv2 using 62 nasopharyngeal swabs and compared the results to RT-qPCR. We created a lyophilized version of ENHANCEv2 and characterized its detection capability and stability.<h4>Results</h4>Here we demonstrate that when coupled with an RT-LAMP step, ENHANCE detects COVID-19 samples down to a few copies with 95% accuracy while maintaining a high specificity towards various isolates of SARS-CoV-2 against 31 highly similar and common respiratory pathogens. ENHANCE works robustly in a wide range of magnesium concentrations (3 mM-13 mM), allowing for further assay optimization. Our clinical validation results for both ENHANCE and ENHANCEv2 show 60/62 (96.7%) sample agreement with RT-qPCR results while only using 5 µL of sample and 20 minutes of CRISPR reaction. We show that the lateral flow assay using paper-based strips displays 100% agreement with the fluorescence-based reporter assay during clinical validation. Finally, we demonstrate that a lyophilized version of ENHANCEv2 shows high sensitivity and specificity for SARS-CoV-2 detection while reducing the CRISPR reaction time to as low as 3 minutes while maintaining its detection capability for several weeks upon storage at room temperature.<h4>Conclusions</h4>CRISPR-based diagnostic platforms offer many advantages as compared to conventional qPCR-based detection methods. Our work here provides clinical validation of ENHANCE and its improved form ENHANCEv2 for the detection of COVID-19.","dates":{"release":"2022-01-01T00:00:00Z","publication":"2022","modification":"2025-04-21T23:16:11.88Z","creation":"2025-02-19T03:27:25.471Z"},"accession":"S-EPMC9053293","cross_references":{"pubmed":["35603267"],"doi":["10.1038/s43856-021-00066-4"]}}