3D single-molecule super-resolution microscopy with a tilted light sheet

Tilted light sheet microscopy with 3D point spread functions (TILT3D) combines a novel, tilted light sheet illumination strategy with long axial range point spread functions (PSFs) for low-background, 3D super-localization of single molecules as well as 3D super-resolution imaging in thick cells. Be...

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Bibliographic Details
Published in:Nature communications 2018-01, Vol.9 (1), p.123-123, Article 123
Main Authors: Gustavsson, Anna-Karin, Petrov, Petar N., Lee, Maurice Y., Shechtman, Yoav, Moerner, W. E.
Format: Article
Language:English
Subjects:
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639/624/1107/328/2237
639/624/1107/328/2238
Cell Line, Tumor
Coding
Construction standards
Emitters
HeLa Cells
Humanities and Social Sciences
Humans
Image resolution
Imaging, Three-Dimensional - methods
Light
Lighting - methods
Localization
Mammalian cells
Mammals
Medicin och hälsovetenskap
Microscopy
Mitochondria
Mitochondria - ultrastructure
multidisciplinary
Point spread functions
Position (location)
Science
Science (multidisciplinary)
Single Molecule Imaging - instrumentation
Single Molecule Imaging - methods
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Summary:Tilted light sheet microscopy with 3D point spread functions (TILT3D) combines a novel, tilted light sheet illumination strategy with long axial range point spread functions (PSFs) for low-background, 3D super-localization of single molecules as well as 3D super-resolution imaging in thick cells. Because the axial positions of the single emitters are encoded in the shape of each single-molecule image rather than in the position or thickness of the light sheet, the light sheet need not be extremely thin. TILT3D is built upon a standard inverted microscope and has minimal custom parts. The result is simple and flexible 3D super-resolution imaging with tens of nm localization precision throughout thick mammalian cells. We validate TILT3D for 3D super-resolution imaging in mammalian cells by imaging mitochondria and the full nuclear lamina using the double-helix PSF for single-molecule detection and the recently developed tetrapod PSFs for fiducial bead tracking and live axial drift correction. Light-sheet single-molecule 3D super-resolution microscopes can’t image close to a coverslip or may require complex apparatus. Here the authors overcome such limitations using a tilted light sheet strategy with long axial range point spread functions on a standard inverted microscope.
ISSN:2041-1723
2041-1723
DOI:10.1038/s41467-017-02563-4