Leitao, S. M.Navikas, V.Miljkovic, H.Drake, B.Marion, S.Pistoletti Blanchet, G.Chen, K.Mayer, S. F.Keyser, U. F.Kuhn, A.Fantner, G. E.Radenovic, A.2023-07-172023-12-192023-07-172023-06-1910.1038/s41565-023-01412-4https://infoscience.epfl.ch/handle/20.500.14299/199165WOS:001010653400001In current nanopore-based label-free single-molecule sensing technologies, stochastic processes influence the selection of translocating molecule, translocation rate and translocation velocity. As a result, single-molecule translocations are challenging to control both spatially and temporally. Here we present a method using a glass nanopore mounted on a three-dimensional nanopositioner to spatially select molecules, deterministically tethered on a glass surface, for controlled translocations. By controlling the distance between the nanopore and glass surface, we can actively select the region of interest on the molecule and scan it a controlled number of times and at a controlled velocity. Decreasing the velocity and averaging thousands of consecutive readings of the same molecule increases the signal-to-noise ratio by two orders of magnitude compared with free translocations. We demonstrate the method's versatility by assessing DNA-protein complexes, DNA rulers and DNA gaps, achieving down to single-nucleotide gap detection. In single-molecule characterization, the near-infinite re-read capability on the same region of interest has the potential to unlock greater sensing capacity. A nanopore-based method, named scanning ion conductance spectroscopy, provides complete control over the translocation speed and nanopore position along a selected region and can detect a single 3 angstrom gap in a long strand of DNA.Nanoscience & NanotechnologyMaterials Science, MultidisciplinaryScience & Technology - Other TopicsMaterials Sciencetrans locationdnamicroscopyplatformvelocitynoisetext::journal::journal article::research article