Quantitative single-molecule localization microscopy reports on protein numbers in signaling protein complexes

Christos Karathanasis; Tim N. Baldering; Carolin Boeger; Marie-Lena I. E. Harwardt; Yunqing Li; Mark S. Schröder; Nina S. Deußner-Helfmann; Mathilda Glaesmann; Sebastian Malkusch; Marina S. Dietz; Anne-Sophie Hafner; Erin Schuman; Gerhard Hummer; Mike Heilemann

doi:10.1117/12.2550635

13 February 2020 Quantitative single-molecule localization microscopy reports on protein numbers in signaling protein complexes

Christos Karathanasis, Tim N. Baldering, Carolin Boeger, Marie-Lena I. E. Harwardt, Yunqing Li, Mark S. Schröder, Nina S. Deußner-Helfmann, Mathilda Glaesmann, Sebastian Malkusch, Marina S. Dietz, Anne-Sophie Hafner, Erin Schuman, Gerhard Hummer, Mike Heilemann

Author Affiliations +

Proceedings Volume 11246, Single Molecule Spectroscopy and Superresolution Imaging XIII; 112460N (2020) https://doi.org/10.1117/12.2550635
Event: SPIE BiOS, 2020, San Francisco, California, United States

Abstract

Knowledge of how proteins organize into functional complexes is essential to understand their biological function. Optical super-resolution techniques provide the spatial resolution necessary to visualize and to investigate individual protein complexes in the context of their cellular environment. Single-molecule localization microscopy (SMLM) builds on the detection of single fluorophore labels, which next to the generation of high-resolution images provides access to quantitative molecular information. We developed various tools for quantitative SMLM (qSMLM), an imaging method that both super-resolves individual protein clusters and reports on molecular numbers by analyzing the kinetics of single emitter blinking. This method is compatible with both fluorescent proteins and organic fluorophores. With qSMLM, we quantify protein copy numbers in single clusters, and we study how changes in the stoichiometry of protein complexes translates into function.

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Christos Karathanasis, Tim N. Baldering, Carolin Boeger, Marie-Lena I. E. Harwardt, Yunqing Li, Mark S. Schröder, Nina S. Deußner-Helfmann, Mathilda Glaesmann, Sebastian Malkusch, Marina S. Dietz, Anne-Sophie Hafner, Erin Schuman, Gerhard Hummer, and Mike Heilemann "Quantitative single-molecule localization microscopy reports on protein numbers in signaling protein complexes", Proc. SPIE 11246, Single Molecule Spectroscopy and Superresolution Imaging XIII, 112460N (13 February 2020); https://doi.org/10.1117/12.2550635

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