Abstract
We report on a straightforward yet powerful implementation of stimulated emission depletion (STED) fluorescence microscopy providing subdiffraction resolution in the far-field. Utilizing the same super-continuum pulsed laser source both for excitation and STED, this implementation of STED microscopy avoids elaborate preparations of laser pulses and conveniently provides multicolor imaging. Operating at pulse repetition rates around 1 MHz, it also affords reduced photobleaching rates by allowing the fluorophore to relax from excitable metastable dark states involved in photodegradation. The imaging of dense nanoparticles and of the microtubular network of mammalian cells evidences a spatial resolution of 30-50 nm in the focal plane, i.e. by a factor of 8-9 beyond the diffraction barrier.
Highlights
Owing to its molecular specificity and notoriously simple sample preparation, far-field fluorescence microscopy has become one of the most commonly applied techniques in the life sciences
We report on a straightforward yet powerful implementation of stimulated emission depletion (STED) fluorescence microscopy providing subdiffraction resolution in the far-field
The advent of stimulated emission depletion (STED) microscopy [1,2,3,4] demonstrated that the limiting role of diffraction can be fundamentally overcome and that fluorescence microscopy with a resolution up to the macromolecular scale is possible with conventional optics
Summary
Owing to its molecular specificity and notoriously simple sample preparation, far-field fluorescence microscopy has become one of the most commonly applied techniques in the life sciences. Abstract: We report on a straightforward yet powerful implementation of stimulated emission depletion (STED) fluorescence microscopy providing subdiffraction resolution in the far-field.
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