csiLSFM combines light-sheet fluorescence microscopy and coherent Structured illumination for a lateral resolution below 100 nm

Bo Jui Chang, Victor Didier Perez Meza, Ernst H.K. Stelzer

Research output: Contribution to journalArticlepeer-review

48 Scopus citations

Abstract

Light-sheet-based fluorescence microscopy (LSFM) features optical sectioning in the excitation process. It minimizes fluorophore bleaching as well as phototoxic effects and provides a true axial resolution. The detection path resembles properties of conventional fluorescence microscopy. Structured illumination microscopy (SIM) is attractive for superresolution because of its moderate excitation intensity, high acquisition speed, and compatibility with all fluorophores. We introduce SIM to LSFM because the combination pushes the lateral resolution to the physical limit of linear SIM. The instrument requires three objective lenses and relies on methods to control two counterpropagating coherent light sheets that generate excitation patterns in the focal plane of the detection lens. SIM patterns with the finest line spacing in the far field become available along multiple orientations. Flexible control of rotation, frequency, and phase shift of the perfectly modulated light sheet are demonstrated. Images of beads prove a near-isotropic lateral resolution of sub-100 nm. Images of yeast endoplasmic reticulum show that coherent structured illumination (csi) LSFMperforms with physiologically relevant specimens.

Original languageEnglish (US)
Pages (from-to)4869-4874
Number of pages6
JournalProceedings of the National Academy of Sciences of the United States of America
Volume114
Issue number19
DOIs
StatePublished - May 9 2017
Externally publishedYes

Keywords

  • LSFM
  • Light sheet
  • SIM
  • Spim
  • Structured illumination

ASJC Scopus subject areas

  • General

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