DNA nanoswitch barcodes for multiplexed biomarker profiling

Abstract

The detection of molecular biomarkers plays a key role in the clinic, aiding in diagnostics and prognostics, and in the research laboratory, contributing to our basic understanding of diseases. The ability to detect multiple and diverse molecular biomarkers within a single accessible assay would have great utility, providing a more comprehensive picture for clinical evaluation and research, but is a challenge with standard methods. One promising approach is the use of dynamic DNA nanostructures that can respond to molecular biomarkers, which have recently been used in a variety of biosensing strategies. In this work, we report the use of programmable DNA nanoswitches for the multiplexed detection of up to 6 biomarkers within a single pot through the use of a barcoded gel-based readout. We demonstrate the barcoding capability using gene fragments that correspond to 6 different diseases, with each fragment or combination of fragments producing a unique barcode signature. As a defining feature of our method, we show "mixed multiplexing" for simultaneous barcoded detection of different types of biomolecules - DNA, RNA, antibody and protein in a single assay. To demonstrate clinical potential, we show multiplexed detection of a prostate cancer biomarker panel in serum that includes two microRNA sequences and prostate specific antigen (PSA). This strategy holds promise in clinical diagnostics for profiling complex and diverse biomarker panels.