Fast widefield scan provides tunable and uniform illumination optimizing super-resolution microscopy on large fields
| Autor: | Mau, Adrien, Friedl, Karoline, Leterrier, Christophe, Bourg, Nicolas, Lévêque-Fort, Sandrine |
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| Přispěvatelé: | Institut des Sciences Moléculaires d'Orsay (ISMO), Université Paris-Saclay-Centre National de la Recherche Scientifique (CNRS), Abbelight, Institut de neurophysiopathologie (INP), Aix Marseille Université (AMU)-Centre National de la Recherche Scientifique (CNRS) |
| Jazyk: | angličtina |
| Rok vydání: | 2021 |
| Předmět: |
0301 basic medicine
Fluorescence-lifetime imaging microscopy Materials science Light Optical sectioning Science [SDV.NEU.NB]Life Sciences [q-bio]/Neurons and Cognition [q-bio.NC]/Neurobiology General Physics and Astronomy Total internal reflection microscopy Field of view Microtubules 01 natural sciences Article General Biochemistry Genetics and Molecular Biology 010309 optics 03 medical and health sciences Optics Chlorocebus aethiops 0103 physical sciences Microscopy Animals Super-resolution microscopy Aster (genus) Lighting [PHYS]Physics [physics] Multidisciplinary Total internal reflection fluorescence microscope biology business.industry Lasers Optical Imaging Reproducibility of Results General Chemistry biology.organism_classification Single Molecule Imaging 030104 developmental biology Wide-field fluorescence microscopy Microscopy Fluorescence COS Cells business Algorithms |
| Zdroj: | Nature Communications Nature Communications, 2021, 12 (1), ⟨10.1038/s41467-021-23405-4⟩ Nature Communications, Nature Publishing Group, 2021, 12 (1), ⟨10.1038/s41467-021-23405-4⟩ Nature Communications, Vol 12, Iss 1, Pp 1-11 (2021) |
| ISSN: | 2041-1723 |
| DOI: | 10.1038/s41467-021-23405-4⟩ |
| Popis: | Non-uniform illumination limits quantitative analyses of fluorescence imaging techniques. In particular, single molecule localization microscopy (SMLM) relies on high irradiances, but conventional Gaussian-shaped laser illumination restricts the usable field of view to around 40 µm × 40 µm. We present Adaptable Scanning for Tunable Excitation Regions (ASTER), a versatile illumination technique that generates uniform and adaptable illumination. ASTER is also highly compatible with optical sectioning techniques such as total internal reflection fluorescence (TIRF). For SMLM, ASTER delivers homogeneous blinking kinetics at reasonable laser power over fields-of-view up to 200 µm × 200 µm. We demonstrate that ASTER improves clustering analysis and nanoscopic size measurements by imaging nanorulers, microtubules and clathrin-coated pits in COS-7 cells, and β2-spectrin in neurons. ASTER’s sharp and quantitative illumination paves the way for high-throughput quantification of biological structures and processes in classical and super-resolution fluorescence microscopies. Uniform illumination is a prerequisite for quantitative analyses in both classical fluorescence microscopy and single molecule localisation microscopy. Here, the authors introduce ASTER, an illumination technique that generates uniform illumination over large and adaptable fields of view, compatible with epifluorescence, HiLo and TIRF illumination schemes. |
| Databáze: | OpenAIRE |
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