Regensburg 2025 – scientific programme

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BP: Fachverband Biologische Physik

BP 7: Single Molecule Biophysics

BP 7.2: Talk

Monday, March 17, 2025, 15:30–15:45, H44

Doubling the resolution of fluorescence-lifetime single-molecule localization microscopy with image scanning microscopy — •Niels Radmacher¹, Oleksii Nevskyi¹, José Ignacio Gallea¹, Jan Christoph Thiele², Ingo Gregor¹, Silvio O. Rizzoli^3,4, and Jörg Enderlein^1,4 — ¹Third Institute of Physics, Georg August University, Göttingen, Germany — ²Department of Chemistry, University of Oxford, Oxford, UK — ³Department of Neuro- and Sensory Physiology, University Medical Center Göttingen, Göttingen, Germany — ⁴Cluster of Excellence - Multiscale Bioimaging: from Molecular Machines to Networks of Excitable Cells (MBExC),Göttingen, Germany

In this study, we integrate a single-photon detector array into a confocal laser scanning microscope, enabling the combination of fluorescence-lifetime single-molecule localization microscopy with image scanning microscopy. This unique combination delivers a twofold improvement in lateral localization accuracy for single-molecule localization microscopy (SMLM) and maintains its simplicity. Moreover, the addition of lifetime information from our confocal laser scanning microscope eliminates chromatic aberration, particularly crucial for achieving few-nanometre resolution in SMLM. Our approach is named fluorescence-lifetime image scanning microscopy iSMLM. And is demonstrated through dSTORM and DNA PAINT experiments on fluorescently labelled cells, showcasing both resolution enhancement and fluorescence-lifetime multiplexing capabilities.

Keywords: super-resolution microscopy; single-molecule localization microscopy; image-scanning microscopy; fluorescence lifetime microscopy