Single-Molecule Detection Concepts Enabled by DNA Origami

Since its introduction in 2006, DNA origami has enabled the fabrication of a wide variety of two- and three-dimensional DNA nanostructures. From the very beginning, researchers have explored these nanostructures as programmable nanobreadboards with hundreds of uniquely addressable positions, allowing precise spatial arrangement of biomolecules, fluorophores, and nanoparticles. This capability has been leveraged to create functional DNA nanomachines capable of single-molecule detection. Here, DNA origami is utilized to precisely engineer various nanoarchitectures, such as conformational switches and plasmonic hotspots. Through coupling of these concepts with tailored readout strategies, true single-molecule detection can be achieved. This literature review systematically examines the development of DNA origami-based single-molecule detection concepts. We first explore general design principles to produce functional DNA nanostructures, followed by an overview of non-fluorescence-based approaches employing atomic force microscopy, nanopores, and optical nanoantennas with surface-enhanced Raman spectroscopy readout, as well as fluorescence-based approaches relying on dynamic DNA nanostructures and optical nanoantennas with fluorescent readout. We highlight key trends as well as the remaining technology gaps that should be bridged to further advance DNA origami towards next-generation single-molecule detection.