Fluorescence Microscopy with 6 nm Resolution on DNA Origami

Resolution of emerging superresolution microscopy is commonly characterized by the width of a point‐spread‐function or by the localization accuracy of single molecules. In contrast, resolution is defined as the ability to separate two objects. Recently, DNA origamis have been proven as valuable scaffold for self‐assembled nanorulers in superresolution microscopy. Here, we use DNA origami nanorulers to overcome the discrepancy of localizing single objects and separating two objects by resolving two docking sites at distances of 18, 12, and 6 nm by using the superresolution technique DNA PAINT(point accumulation for imaging in nanoscale topography). For the smallest distances, we reveal the influence of localization noise on the yield of resolvable structures that we rationalize by Monte Carlo simulations.