GSD microscopy
Encyclopedia
Ground State Depletion Microscopy, or GSD Microscopy, is an implementation of the RESOLFT
concept. The method was proposed in 1995 and experimentally demonstrated in 2007. It is the second concept to overcome the diffraction barrier in far-field optical microscopy published by Stefan Hell
. Using nitrogen-vacancy center
s in diamonds a resolution of up to 7.8 nm was achieved in 2009. This is far below the diffraction limit (~200 nm).
), it cannot be excited from the ground state. Switching between these two states (bright and dark) by applying light fulfills all preconditions for the RESOLFT
concept and subdiffraction imaging and images with very high resolution can be obtained. For successful realization, GSD microscopy requires either special fluorophores with high triplet yield, or removal of oxygen by use of various mounting medium like Moviol or Vectashield.
The implementation in a microscope is very similar to Stimulated Emission Depletion Microscopy
, however it can operate with only one wavelength for excitation and depletion. Using an appropriate ring-like focal spot for the light that brings the molecules into the dark state, the fluorescence can be quenched at the outer part of the focal spot. Therefore fluorescence only still takes place at the center of the microscope's focal spot and the spatial resolution is increased.
RESOLFT
RESOLFT, an acronym for REversible Saturable OpticaL Fluorescence Transitions, denotes a group of optical microscopy techniques with very high resolution...
concept. The method was proposed in 1995 and experimentally demonstrated in 2007. It is the second concept to overcome the diffraction barrier in far-field optical microscopy published by Stefan Hell
Stefan Hell
Stefan W. Hell is a physicist and one of the directors of the Max Planck Institute for Biophysical Chemistry in Göttingen, Germany as well as the head of the department "Optical Nanoscopy" at the German Cancer Research Center in Heidelberg.- Life :In 1981 Hell began his studies at the...
. Using nitrogen-vacancy center
Nitrogen-vacancy center
The nitrogen-vacancy center is one of numerous point defects in diamond. Its most explored and useful property is photoluminescence, which can be easily detected from an individual N-V center...
s in diamonds a resolution of up to 7.8 nm was achieved in 2009. This is far below the diffraction limit (~200 nm).
Principle
In GSD microscopy fluorescent markers are used. In one condition the marker can freely be excited from ground state and returns spontaneously via emission of a fluorescence photon. However if light of appropriate wavelength is additionally applied the dye can be excited to a long-lived dark state, i.e. a state where no fluorescence occurs. As long as the molecule is in the long-lived dark state (e.g. a triplet stateTriplet state
A spin triplet is a set of three quantum states of a system, each with total spin S = 1 . The system could consist of a single elementary massive spin 1 particle such as a W or Z boson, or be some multiparticle state with total spin angular momentum of one.In physics, spin is the angular momentum...
), it cannot be excited from the ground state. Switching between these two states (bright and dark) by applying light fulfills all preconditions for the RESOLFT
RESOLFT
RESOLFT, an acronym for REversible Saturable OpticaL Fluorescence Transitions, denotes a group of optical microscopy techniques with very high resolution...
concept and subdiffraction imaging and images with very high resolution can be obtained. For successful realization, GSD microscopy requires either special fluorophores with high triplet yield, or removal of oxygen by use of various mounting medium like Moviol or Vectashield.
The implementation in a microscope is very similar to Stimulated Emission Depletion Microscopy
STED microscopy
Stimulated Emission Depletion microscopy, or STED microscopy, is a fluorescence microscopy technique that uses the non-linear de-excitation of fluorescent dyes to overcome the resolution limit imposed by diffraction with standard confocal laser scanning microscopes and conventional far-field...
, however it can operate with only one wavelength for excitation and depletion. Using an appropriate ring-like focal spot for the light that brings the molecules into the dark state, the fluorescence can be quenched at the outer part of the focal spot. Therefore fluorescence only still takes place at the center of the microscope's focal spot and the spatial resolution is increased.
See also
- Department of NanoBiophotonics at the Max-Planck-Institute for Biophysical Chemistry (Göttingen, Germany)