Mikroskopie

We present a high resolution fluorescence microscope setup which combines the advantages of Structured Illumination Microscopy (SIM) with those of Single Molecule Localization Microscopy (SMLM) in a single instrument. While localization microscopy methods provide highly accurate (10~20nm) position information of optically isolated targets down to the nanometer range, SMLM-data typically consists of less fluorophores contributing to an image compared to a SIM-acquisition. Furthermore, visualization algorithms applied on SMLM data often introduce various artifacts, which do not occur in SIM-images. A combination of the SMLM-data with a wide-field SIM-image revealing the structure allows assigning position information directly to the highly resolved (~100nm) structural information of the specimen acquired with SIM and allows identifying artifacts produced during visualization processing. The same regions of interest can easily be imaged in the two microscope modes, and switching between the two modes is done quickly. We describe the combined setup and its control software together with first applications demonstrating its unique advantages. Several additional features of the versatile SIM-SMLM-microscope including a user-friendly fully automated and time-saving acquisition script are presented, making this combination of two high-resolution techniques superior over existing solutions.