CAREER: Ultrasensitive Multiplex Nanopore Biosensing for Point-of-Care Testing of Infectious Diseases

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The ongoing global pandemic of COVID-19 has reminded us the importance of diagnostic testing technologies. Current point-of-care testing (POCT) technologies are inexpensive and easy to use, store, and transport. However, their simplicity causes a significant loss of sensitivity and specificity comparing to lab based diagnostic tests. The major goal of this NSF CAREER research program is to develop a new nanopore testing technology to enable ultrasensitive detection of infectious diseases at a reasonably low cost. Upon the successful completion of this project, the proposed nanopore test will meet the urgent need for a POCT technology with accuracy that exceeds current POCTs or even lab-based testing technologies. This platform technology presents great potential for changing the current paradigm of POCT for existing infectious diseases. Also, it can be readily modified with minimal optimization as soon as new diseases and biomarkers are identified for rapid deployment in clinics and at the point of care. This project also includes education and outreach activities, such as organizing a summer research workshop and internship for high school students on the fundamentals of Biomedical Engineering and Nanotechnology, broadly presenting our research to local communities, and starting an infectious disease research symposium that brings an additional synergy among local clinicians, biomedical engineers, and scientists. Special attention will be given to students from underrepresented groups and first-generation college students, who constitute a significant part of our student population. This CAREER program should result in achievement awards and authorships on research publications for undergraduate and high school students.In the era of personalized medicine, rapid and accurate quantification of multiple biomarkers at the point-of-care is fundamental to a successful control and management of infectious disease outbreaks. Classical point-of-care testing (POCT) technologies are inexpensive and easy to use, store, and transport. But their simplicity causes a significant loss of sensitivity and specificity comparing to lab based in vitro diagnostics. The nanopore technology is a promising alternative because of its single-molecule analysis capacity, portability, and low cost. However, existing nanopore technologies are not suitable for detecting analytes in complex human samples, can only analyze charged biomarkers of certain sizes, and are not device compatible. The major goal of this CAREER program is to develop a next generation nanopore biosensor to enable ultrasensitive detection and quantification of multiple infectious disease biomarkers from human blood at the point of care. The program has three research objectives: (1) Develop and optimize a nanopore biosensor with unprecedented sensitivity, robustness, and reproducibility for various purposes; (2) Enable sensitive and specific biomarker detection by automated immunoprecipitation and novel signal transduction mechanisms; (3) Demonstrate ultrasensitive multiplex quantification of circulating proteomics biomarkers and its applications in infectious disease diagnosis and prognosis. Proposed approach has several innovative elements: several amplification methods are employed to achieve the sub-femtomolar level detection limit; special single strand DNA structures are used as detection surrogates for biomarkers, so that the readout signal can be easily differentiated from other biomolecules to prevent false-positives; an integrated microfluidic automatic immunoprecipitation module can reduce sample-to-answer time and human errors in the assay protocol.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.