EAGER SENTINELS: The PCR-free Biosensor for a Fast, Simple, and Sensitive Detection of RNA.

The recent pandemic has highlighted the urgent need for rapid and accurate detection of viruses to allow for early disease diagnosis and monitoring to prevent future pandemics and to reduce the risk of complications and mortality through timely health care. Currently, diagnostic tests based on polymerase chain reaction (PCR) are widely applied for the detection of viruses. Despite outstanding analytical parameters, significant drawbacks of this technology have become evident during the recent COVID pandemic. More specifically, PCR-based tests require not only expensive laboratory equipment and highly trained personnel, but they are also time-consuming and not well adapted for point-of-care devices. For example, it usually takes two to three days to get the result of a PCR-based COVID-19 test. Consequently, the spread of the virus becomes less containable. The purpose of this project is to develop a fast, easy, and economically feasible biosensing platform that does not require PCR. This project also aims to provide undergraduate students with interdisciplinary training in the development of biosensors, and local high school students with an opportunity to explore the field of diagnostics. This project is to design and realize a rapid, ultrasensitive, and adaptable PCR-free biosensing platform for viral RNA detection. Specifically, the proposed biosensor will be based on the newly discovered CRISPR-Cas (Clustered Regularly Interspaced Short Palindromic Repeat – CRISPR associated) nuclease that shows high selectivity, and can be reprogrammed to detect various viruses’ RNAs. The development and integrations of a novel signal amplification scheme will allow circumvention of the PCR amplification step. Not only will this research lead to the novel RNA sensing approach, but it will also provide a blueprint for developing a DNA biosensor based on an alternative but closely related Cas nuclease. Due to the ease of configuration and operation, the proposed biosensor is more economical, faster, and will not require sophisticated equipment and personnel training, thereby addressing current needs in public health. This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.