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Abstract:
We report on a fiber laser-based stimulated emission-depletion microscope providing down to ~20 nm resolution
in raw data images as well as 15–19 nm diameter probing areas in fluorescence correlation spectroscopy. Stimulated emission
depletion pulses of nanosecond duration and 775 nm wavelength are used to silence two fluorophores simultaneously, ensuring
offset-free colocalization analysis. The versatility of this superresolution method is exemplified by revealing the octameric
arrangement of Xenopus nuclear pore complexes and by quantifying the diffusion of labeled lipid molecules in artificial and living
cell membranes.