Regensburg 2022 – wissenschaftliches Programm

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BP: Fachverband Biologische Physik

BP 5: Focus Session: Super Resolution Microscopy and Dynamics of Supramolecular Complexes

BP 5.2: Vortrag

Montag, 5. September 2022, 15:30–15:45, H15

Photon-stream-based aberration correction for STED microscopy — •Debadrita Ghosh, Claudia Geisler, and Alexander Egner — Institute for Nanophotonics, Goettingen, Germany

Stimulated emission depletion (STED) microscopy is the most prominent super-resolution fluorescence microscopy method and achieves a resolution far beyond the diffraction limit. However, like all these methods, it is adversely affected by sample-induced aberrations, which can degrade the achievable resolution and image quality significantly. These aberrations are caused by wavefront distortions due to refractive index variations, for example within thick biological specimens. This challenge can be addressed by using adaptive optics (AO) in a feedback-controlled manner such that the wavefront distortions are compensated and the image quality is restored. Typically, the feed-back loop uses image features which necessitates repeated acquisitions of the same field-of-view. This approach, therefore, is slow and prone to unwanted photo-bleaching. Here, we present an AO correction scheme that does not rely on image features, but exploits the dependence of the fluorescence lifetime on the local STED intensity. In principle, our photon-stream-based metric can be evaluated on a single image pixel, which makes it photon-budget-friendly and allows to correct aberrations rapidly in parallel with image acquisition. We successfully utilized this new metric for automated and continuous aberration correction in biological samples, making imaging fast and easy even for users without expert knowledge.