Super-localization of individual fluorophores along a DNA strand in a microchannel

Abstract

DNA mapping is a method of stretching random-coiled DNA molecules and then analyzing them using a fluorescence microscope. This method has been used for DNA analyses. In this study, to realize more accurate DNA analyses with small amounts of samples, we aimed to stretch and immobilize λ DNA molecules and to achieve super-resolution imaging with the direct stochastic optical reconstruction microscopy (dSTORM) of a single λ DNA molecule in a microchannel. To stretch and immobilize the DNA molecule, we used an air–water interface movement by controlling the pressure in the microchannel. The DNA molecule was stretched and immobilized on an air-plasma-treated glass substrate, which prevented the overlapping of the DNA molecules owing to the small adhesion force, and a stretching ratio of 75% was achieved. We performed dSTORM imaging with the blinking of YOYO-1 dyes along the DNA molecule in the microchannel with the width of 200 μm, the depth of 2 μm, and the length of 40 mm. We obtained the super-resolution imaging of the DNA molecule with the full width at half maximum of 67 nm. The design of microchannel is required to improve dSTORM imaging of DNA molecules, and the issue could be explored in our future studies.