Abstract
Single-molecule localization microscopy (SMLM) has become a popular tool in cell biology as it allows imaging cellular structures with a precision of tens of nanometers. However, acquiring superresolved images with SMLM is inherently slow, requiring up to hours for a single image and careful user intervention in between experiments. This effectively limits the throughput of SMLM to a handful of images of the structure of interest per study. We present tools to overcome this limitation by automating a wide-field microscope to perform SMLM experiments. This microscope is capable of SMLM imaging over the course of days thanks to advanced electronics and an open-source plugin for Micro-manager. In the meantime, SMLM experiments are analysed during acquisition using a freely accessible software in Matlab. Finally, the microscope features a flat illumination system, providing homogeneous statistics across large fields of view. Altogether these developments in optics, hardware and software can be transferred to any single-molecule wide-field microscope, thus enabling automated localization microscopy.
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