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A quantitative analysis of various patterns applied in lattice light sheet microscopy

Author

Listed:
  • Yu Shi

    (University of North Carolina at Chapel Hill, North Carolina State University)

  • Timothy A. Daugird

    (University of North Carolina at Chapel Hill)

  • Wesley R. Legant

    (University of North Carolina at Chapel Hill, North Carolina State University
    University of North Carolina at Chapel Hill)

Abstract

Light sheet microscopes reduce phototoxicity and background and improve imaging speed compared to widefield and confocal microscopes. However, when equipped with Gaussian beams, the axial resolving power of a light sheet microscope and the observable field of view are inversely related. Light sheets based on dithered optical lattices improve axial resolution and beam uniformity compared Gaussian beams by using axially structured illumination patterns. However, these advantages come at the expense of an increased total illumination to the specimen and a decreased axial confinement of the illumination pattern. Using simulations and experimental measurements in fixed and live cells, we quantify the differences between Gaussian and lattice light sheets on beam uniformity, axial resolution, lateral resolution, and photobleaching. We demonstrate how different optical lattice illumination patterns can be tuned to prioritize either axial resolution or optical sectioning. Finally, we introduce an approach to spectrally fuse sequential acquisitions of different lattice light sheet patterns with complementary optical properties to achieve both high resolution and low background images.

Suggested Citation

  • Yu Shi & Timothy A. Daugird & Wesley R. Legant, 2022. "A quantitative analysis of various patterns applied in lattice light sheet microscopy," Nature Communications, Nature, vol. 13(1), pages 1-12, December.
    • Handle: RePEc:nat:natcom:v:13:y:2022:i:1:d:10.1038_s41467-022-32341-w
    DOI: 10.1038/s41467-022-32341-w
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    Cited by:

    1. Timothy A. Daugird & Yu Shi & Katie L. Holland & Hosein Rostamian & Zhe Liu & Luke D. Lavis & Joseph Rodriguez & Brian D. Strahl & Wesley R. Legant, 2024. "Correlative single molecule lattice light sheet imaging reveals the dynamic relationship between nucleosomes and the local chromatin environment," Nature Communications, Nature, vol. 15(1), pages 1-20, December.

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