Nature Methods
- 4, 915 - 918 (2007)
Published online: 21 October 2007; | doi:10.1038/nmeth1108
STED microscopy with continuous wave beamsKatrin I Willig1, 2, Benjamin Harke1, 2, Rebecca Medda1 & Stefan W Hell11
Max Planck Institute for Biophysical Chemistry, Department of NanoBiophotonics, Am Fassberg 11, 37077 Göttingen, Germany. 2
These authors contributed equally to this work.
Correspondence should be addressed to Stefan W Hell shell@gwdg.de 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.
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