Rapid and Fully Microfluidic Ebola Virus Detection with CRISPR-Cas13a

Ebola virus causes a fatal hemorrhagic fever, which is zoonotic and endemic in some developing countries. Several deadly outbreaks have been recorded in the past, and timely diagnosis is always critical. Immunoassays that have been developed lack sufficient sensitivity and although PCR-based systems are the current gold standard, their use is limited by the need for high expertise, expensive reagents and sophisticated instrumentation. This creates need for a simple, cost effective and highly sensitive diagnostic method. In this study a technology based on the CRISPR-Cas13 enzyme is employed. The enzyme is bound to a guide RNA sequence that is complementary to the target viral RNA. When the target RNA binds, the enzyme is activated and begins to cleave the RNA molecules. A fluorescent RNA probe tagged with a fluorophore and a quencher is included, when the probe is cut the fluorophore is released and emits a detectable signal. The intensity of the signal is measured and quantified. Key advantages of this method are that it does not require RNA amplification and provides high sensitivity and specificity. In addition, the technology uses a microfluidic chip to automate reagent mixing and incubation, reducing the total detection time about five minutes. 
(SWM)