STED microscopy with continuous wave beams

Nat Methods. 2007 Nov;4(11):915-8. doi: 10.1038/nmeth1108. Epub 2007 Oct 21.

Abstract

We report stimulated emission depletion (STED) fluorescence microscopy with continuous wave (CW) laser beams. Lateral fluorescence confinement from the scanning focal spot delivered a resolution of 29-60 nm in the focal plane, corresponding to a 5-8-fold improvement over the diffraction barrier. Axial spot confinement increased the axial resolution by 3.5-fold. We observed three-dimensional (3D) subdiffraction resolution in 3D image stacks. Viable for fluorophores with low triplet yield, the use of CW light sources greatly simplifies the implementation of this concept of far-field fluorescence nanoscopy.

MeSH terms

  • Animals
  • Cell Line, Tumor
  • Fluorescent Dyes / chemistry
  • Humans
  • Imaging, Three-Dimensional
  • Immunohistochemistry
  • Lamins / analysis
  • Lasers*
  • Microscopy, Confocal
  • Microscopy, Fluorescence / instrumentation
  • Microscopy, Fluorescence / methods*
  • Microspheres
  • Neurofilament Proteins / analysis
  • Nuclear Lamina / chemistry
  • Nuclear Lamina / metabolism
  • Optics and Photonics*
  • PC12 Cells
  • Qa-SNARE Proteins / analysis
  • Rats
  • Spectrometry, Fluorescence

Substances

  • Fluorescent Dyes
  • Lamins
  • Neurofilament Proteins
  • Qa-SNARE Proteins