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Lateral resolution of 28 nm (lambda/25) in far-field fluorescence microscopy
No full textWe demonstrate sub-diffraction lateral resolution of 28 +/- 2 nm in far-field fluorescence microscopy through stimulated emission depletion effected by an amplified laser diode. Measurement of the optical transfer function in the focal plane reveals a 6-fold enlargement of the spatial bandwith over the diffraction limit. The resolution is established by imaging individual fluorescent molecules on a surface. Corresponding to 1/25 of the responsible wavelength, the attained resolution represents a new benchmark in far-field microscopy and underscores the viability of fluorescence nanoscopy with visible light, conventional optics and compact laser systems
Absolute optical cross section of individual fluorescent molecules
No full textLight switches: The photon capture area of single fluorescent molecules has been directly measured by switching them “on and off” with light. Stimulated emission of single molecules (see picture) enables the precise control of the excited state population probability of an individual molecule at room temperature
Fluoreszenzmikroskopie sieht chemisch scharf
No full textObwohl das Stimulated‐Emission‐Depletion (STED)‐Fluoreszenzmikroskop Licht auf herkömmliche Weise fokussiert, ist die Auflösung nicht durch Beugung begrenzt und kann prinzipiell molekulare Dimensionen erreichen. Die zugrunde liegende Idee gilt für alle reversibel schaltbaren optischen Übergänge eines Markermoleküls. So wäre die Beugungsgrenze für optisches Abbilden und Strukturieren überwunden
Absolute optische Wirkungsquerschnitte fluoreszierender Einzelmoleküle
No full textLichtschalter: Der Photoneneinfangquerschnitt einzelner fluoreszierender Moleküle ist direkt messbar, indem man angeregte Moleküle mit Licht „ausschaltet“. Diese stimulierte Emission (siehe Abbildung) ermöglicht die direkte Manipulation des angeregten Zustandes eines einzelnen Moleküls bei Raumtemperatur
Imaging and writing at the nanoscale with focused visible light through saturable optical transitions
No full textEffecting a saturable optical molecular transition with a spatial intensity distribution featuring a local minimum allows the fundamental breaking of the diffraction barrier both in microscopy and in material structuring. If the transition can be repeatedly reverted, as in switchable fluorescent proteins and photochromic compounds, fluorescence imaging and writing is possible with spatial resolution down to the molecular scale
STED microscopy with a supercontinuum laser source.
No full textWe 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
A compact STED microscope providing 3D nanoscale resolution.
No full textThe advent of supercontinuum laser sources has enabled the implementation of compact and tunable stimulated emission depletion fluorescence microscopes for imaging far below the diffraction barrier. Here we report on an enhanced version of this approach displaying an all-physics based resolution down to (19 +/- 3) nm in the focal plane. Alternatively, this single objective lens system can be configured for 3D imaging with resolution down to 45 x 45 x 108 nm in a cell. The obtained results can be further improved by mathematical restoration algorithms. The far-field optical nanoscale resolution is attained in a variety of biological samples featuring strong variations in the local density of features
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