High speed optical nanoscopy by stimulated emission depletion (STED) with galvo mirrors

摘要

Stimulated emission depletion microscopy (STED) has been proved to be a feasible and straightforward method of breaking the conventional diffraction barrier in the far-field. In this paper, we design and setup a home-built high speed STED microscope. Both the excitation beam (488nm) and the depletion beam (592nm) are provided by continuous wave lasers. By using a pair of galvo mirrors (TILL Yanus IV Digital Scan Head), we realize a pixel dwell time down to 2 μs in the experiment which enables an acquisition speed of 2 frames per second in an imaging field of 5*5 μm with an individual pixel size of 10nm. The image acquisition process is controlled by the software Imspector. In the manuscript, we give a clear description on how to build the microscope and also conduct several experiments to evaluate its performance in practice. A spatial resolution of ＜100nm, which is well beyond the diffraction barrier has been demonstrated in both nanoparticles and biomedical samples. Featuring a superresolution together with a high imaging speed, our STED microscope has big potential to be widely applied in related scientific researches.