Visual detection of SARS-CoV-2 with a CRISPR/Cas12b-based platform

Abstract

The coronavirus disease 2019 (COVID-19) caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has caused a global pandemic, highlighting the unprecedented demand for rapid and portable diagnostic methods. Clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated (Cas) proteins-based platforms have been used for the detection of pathogens. However, in further applications and research, due to multiple steps needed, many methods showed an increased risk of cross-reactivity. The thermostable Cas12b enables the combination of isothermal amplification and CRISPR-mediated detection, which could decrease the risk of cross-contamination. In this study, we developed a portable and specific diagnostic method that combined the gold nanoparticle (AuNP) with thermal stable CRISPR/Cas12b-enhanced reverse transcription loop-mediated isothermal amplification (RT-LAMP), which is called SCAN, to distinguish the N gene of SARS-CoV-2 from flu gene. We validated our method using RNA from cells transfected by plasmids. We could easily distinguish the positive results by the naked eye based on the strong molar absorption coefficient of AuNP. Moreover, SCAN has the potential for high-throughput tests owing to its convenient operation. In sum, SCAN has broken the site and equipment restrictions of traditional detection methods and could be applied outside of hospitals and clinical laboratories, greatly expanding the test of COVID-19.