Multiplexed electronic counting of scarce protein targets using nucleic acid nanoparticles

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PROJECT SUMMARY This interdisciplinary project synergizes the expertise of three research groups for a proof-of-principle demonstration of a novel approach for accurate and sensitive detection of scarce protein biomarkers. The key innovative element of the approach is the use of wireframe-like nucleic acid nanoparticles (NANPs) to bind protein targets with high affinity either directly or by means of auxiliary antibody proteins. Binding of the protein targets is detected by first incubating the sample with a cocktail of NANPs and then examining them using a nanopore in a solid-state membrane. Protein detection relies on measurement of the ionic current flowing through the nanopore: the ionic current and particle dwell time decreases by a characteristic amount when a NANP of a certain type enters the nanopore. Importantly, by matching the physical dimensions of the NANP to the physical dimension of the nanopore, we expect to dramatically increase the residence type of the nucleic acid nanoparticles within the nanopore and thereby achieve ultra-sensitive (sub-picomolar range) detection of the protein-bound biomarkers. By designing our NANP probes to produce distinct ionic signatures when bound to their protein targets, we will achieve multiplex detection of several protein species using the same nanopore as well as combinatorial detection of biomarkers by assembling the NANP probes and protein biomarkers into a sandwich like structures. The project will be carried out by the Afonin group at UNC Charlotte that will design NANPs, the Wanunu group at Northeastern University that will perform the nanopore detection experiments, and the Aksimentiev group that will use an arsenal of modeling techniques to optimize and improve the detection strategy. Our ultra-sensitive, portable, rapid, and potentially low-cost technology for quantification of protein levels is expected to find broad use for the analysis of biological samples, eventually offering sensitive, reliable, and minimally invasive identification of disease-indicative biomarkers that could be important innovations for early-stage diagnostics of cancer and other diseases.