CRISPR Microfluidic Array for Simultaneous Multiplexed Sensing of Proteins and miRNAs

University Of Connecticut, Storrs CT

Investigators

Abstract

Measurement of biomolecules like proteins and nucleic acids, called biomarkers, in blood, is a valuable approach to detect cancers and other diseases. Samples consisting of several proteins and nucleic acids show promise for this purpose, but at present must be measured using separate, time-consuming methods. This project uses a biological construct called CRISPR (Clustered Regularly Interspaced Short Palindromic Repeats) to enable measurements. CRISPR can be designed to bind a nucleic acid or a protein labeled with a nucleic acid and give a signal only when that specific nucleic acid or label binds to CRISPR. This approach will be developed into a fast, low-cost 3D-printed, semi-automated device to measure several nucleic acids and several protein biomarkers in the same test. The project will also develop sensitive electrically-driven light emission called electrochemiluminescence (ECL) as a simpler detection method. The blood test using the 3D-printed device will be designed to run in a short time in a clinic or physician’s office by a lightly trained technician. This will allow fast turnaround in what is called point of care mode. Results will be available to the physician while the patient is still in his or her office and can be used at that time as part of the diagnosis. This project will utilize programs at Univ. of Connecticut to support education in new CRISPR technology for graduate students, undergraduates, science teachers and high school students. Activities include holding workshops on CRISPR detection through SECRET (School of Exploratory Chemistry Research Experience and Training), developing Early College Experience programs for secondary school students and science teachers, facilitating training of Ph.D. students in CRISPR array design and fabrication, and providing research opportunities for undergraduates at UCONN and in UCONN’s NSF REU program, as well as high school students through the ACS SEED program. Molecular biomarkers have become increasingly valuable in medical diagnostics for many diseases. Panels of biomarkers, as opposed to single biomarkers, show superior diagnostic utility and combined protein-miRNA panels have shown significantly improved diagnostic predictions over separate protein or miRNA panels. This project will develop the first analytical array using CRISPR technology to simultaneously measure combinations of miRNAs and proteins. The new 3D-printed array will be able to analyze miRNA-protein biomarker panels for a wide range of disease diagnostics in point-of-care (POC) mode, as well as in other biomedical research applications. In addition to fluorescent detection, electrochemiluminescence (ECL) will be explored with CDC and iPhone camera detection. The focus will be on miRNA and protein biomarkers that are high performing biomarkers for aggressive prostate cancer. Assays will be validated by measuring target miRNAs and proteins in 130 existing human serum samples from patients with high grade prostate cancers and controls. The intellectual merit of this project involves design and development of the first engineering strategy using a new CRISPR assay technology to detect panels of miRNAs and protein simultaneously at POC. 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.