Rapid and Fully Microfluidic Ebola Virus Detection with CRISPR-Cas13a

作者：Peiwu Qin, Myeongkee Park, Kendra J. Alfson, Manasi Tamhankar, Ricardo Carrion, Jean L. Patterson, Anthony Griffiths, Qian He, Ahmet Yıldız, Richard A. Mathies, Ke Du · 发表于：ACS Sensors · 年份：2019 · DOI：10.1021/acssensors.9b00239 · 被引用次数：325 · 研究领域：CRISPR and Genetic Engineering、Viral Infections and Outbreaks Research、Viral gastroenteritis research and epidemiology

Highly infectious illness caused by pathogens is endemic especially in developing nations where there is limited laboratory infrastructure and trained personnel. Rapid point-of-care (POC) serological assays with minimal sample manipulation and low cost are desired in clinical practice. In this study, we report an automated POC system for Ebola RNA detection with RNA-guided RNA endonuclease Cas13a, utilizing its collateral RNA degradation after its activation. After automated microfluidic mixing and hybridization, nonspecific cleavage products of Cas13a are immediately measured by a custom integrated fluorometer which is small in size and convenient for in-field diagnosis. Within 5 min, a detection limit of 20 pfu/mL (5.45 × 10 7 copies/mL) of purified Ebola RNA is achieved. This isothermal and fully solution-based diagnostic method is rapid, amplification-free, simple, and sensitive, thus establishing a key technology toward a useful POC diagnostic platform.