Limits...
Super-resolution molecular and functional imaging of nanoscale architectures in life and materials science.

Habuchi S - Front Bioeng Biotechnol (2014)

Bottom Line: Super-resolution (SR) fluorescence microscopy has been revolutionizing the way in which we investigate the structures, dynamics, and functions of a wide range of nanoscale systems.I discuss the applications of SR microscopy in the fields of life science and materials science with a special emphasis on quantitative molecular imaging and nanoscale functional imaging.These studies open new opportunities for unraveling the physical, chemical, and optical properties of a wide range of nanoscale architectures together with their nanostructures and will enable the development of new (bio-)nanotechnology.

View Article: PubMed Central - PubMed

Affiliation: Biological and Environmental Sciences and Engineering Division, King Abdullah University of Science and Technology , Jeddah , Saudi Arabia.

ABSTRACT
Super-resolution (SR) fluorescence microscopy has been revolutionizing the way in which we investigate the structures, dynamics, and functions of a wide range of nanoscale systems. In this review, I describe the current state of various SR fluorescence microscopy techniques along with the latest developments of fluorophores and labeling for the SR microscopy. I discuss the applications of SR microscopy in the fields of life science and materials science with a special emphasis on quantitative molecular imaging and nanoscale functional imaging. These studies open new opportunities for unraveling the physical, chemical, and optical properties of a wide range of nanoscale architectures together with their nanostructures and will enable the development of new (bio-)nanotechnology.

No MeSH data available.


Principles of super-resolution fluorescence structural imaging. Schematic illustrations of the principles of (A) STED microscopy, (B) (S)SIM, and (C) localization microscopy.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC4126472&req=5

Figure 1: Principles of super-resolution fluorescence structural imaging. Schematic illustrations of the principles of (A) STED microscopy, (B) (S)SIM, and (C) localization microscopy.

Mentions: The concept of stimulated-emission depletion (STED) microscopy was proposed by Stefan Hell (Hell and Wichmann, 1994) and was later demonstrated experimentally (Klar et al., 2000). In STED microscopy, spontaneous fluorescence emission caused by the excitation of the fluorophores by an excitation laser with diffraction-limited spot size is suppressed by a second laser beam (STED beam), which depletes the excited-state through stimulated-emission before the fluorophores emit light through spontaneous fluorescence (Figure 1A). The STED beam with a doughnut-like spatial pattern leads to the depletion of the spontaneous fluorescence at the periphery of the fluorescence spot. When the fluorescence is depleted by an intense STED beam, the fluorescence depletion in the peripheral region is saturated (i.e., saturation of stimulated-emission), resulting in a very small fluorescence spot at the region around the focal point. This corresponds to the reduction of the PSF. An SR fluorescence image can be obtained by scanning the spatially overlapped excitation and STED beams across the sample. Analogous methods such as ground-state depletion (GSD) microscopy (Bretschneider et al., 2007) have been developed using different mechanisms of fluorescence depletion. Collectively, these methods are referred to as reversible saturable optical fluorescence transitions (RESOLFT) microscopy (Hofmann et al., 2005). A spatial resolution of approximately 30 nm in both the lateral and axial directions has been achieved using a STED beam of a 3D doughnut-like spatial pattern (Schmidt et al., 2008). STED microscopy with an imaging speed similar to confocal microcopy (i.e., video-rate) has been reported (Westphal et al., 2008). A rapid RESOLFT microscopy imaging (120 μm × 100 μm-sized fields of view in <1 s) has also been achieved by parallelizing scanning with more than 100,000 beams (Chmyrov et al., 2013). Furthermore, the spatial resolution has been improved by the time-gated detection of fluorescence (Vicidomini et al., 2011), which allows the visualizing of finer structures at relatively low intensities of the STED beam. STED microscopy has also been combined with two-photon excitation microscopy (Bianchini et al., 2012).


Super-resolution molecular and functional imaging of nanoscale architectures in life and materials science.

Habuchi S - Front Bioeng Biotechnol (2014)

Principles of super-resolution fluorescence structural imaging. Schematic illustrations of the principles of (A) STED microscopy, (B) (S)SIM, and (C) localization microscopy.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4126472&req=5

Figure 1: Principles of super-resolution fluorescence structural imaging. Schematic illustrations of the principles of (A) STED microscopy, (B) (S)SIM, and (C) localization microscopy.
Mentions: The concept of stimulated-emission depletion (STED) microscopy was proposed by Stefan Hell (Hell and Wichmann, 1994) and was later demonstrated experimentally (Klar et al., 2000). In STED microscopy, spontaneous fluorescence emission caused by the excitation of the fluorophores by an excitation laser with diffraction-limited spot size is suppressed by a second laser beam (STED beam), which depletes the excited-state through stimulated-emission before the fluorophores emit light through spontaneous fluorescence (Figure 1A). The STED beam with a doughnut-like spatial pattern leads to the depletion of the spontaneous fluorescence at the periphery of the fluorescence spot. When the fluorescence is depleted by an intense STED beam, the fluorescence depletion in the peripheral region is saturated (i.e., saturation of stimulated-emission), resulting in a very small fluorescence spot at the region around the focal point. This corresponds to the reduction of the PSF. An SR fluorescence image can be obtained by scanning the spatially overlapped excitation and STED beams across the sample. Analogous methods such as ground-state depletion (GSD) microscopy (Bretschneider et al., 2007) have been developed using different mechanisms of fluorescence depletion. Collectively, these methods are referred to as reversible saturable optical fluorescence transitions (RESOLFT) microscopy (Hofmann et al., 2005). A spatial resolution of approximately 30 nm in both the lateral and axial directions has been achieved using a STED beam of a 3D doughnut-like spatial pattern (Schmidt et al., 2008). STED microscopy with an imaging speed similar to confocal microcopy (i.e., video-rate) has been reported (Westphal et al., 2008). A rapid RESOLFT microscopy imaging (120 μm × 100 μm-sized fields of view in <1 s) has also been achieved by parallelizing scanning with more than 100,000 beams (Chmyrov et al., 2013). Furthermore, the spatial resolution has been improved by the time-gated detection of fluorescence (Vicidomini et al., 2011), which allows the visualizing of finer structures at relatively low intensities of the STED beam. STED microscopy has also been combined with two-photon excitation microscopy (Bianchini et al., 2012).

Bottom Line: Super-resolution (SR) fluorescence microscopy has been revolutionizing the way in which we investigate the structures, dynamics, and functions of a wide range of nanoscale systems.I discuss the applications of SR microscopy in the fields of life science and materials science with a special emphasis on quantitative molecular imaging and nanoscale functional imaging.These studies open new opportunities for unraveling the physical, chemical, and optical properties of a wide range of nanoscale architectures together with their nanostructures and will enable the development of new (bio-)nanotechnology.

View Article: PubMed Central - PubMed

Affiliation: Biological and Environmental Sciences and Engineering Division, King Abdullah University of Science and Technology , Jeddah , Saudi Arabia.

ABSTRACT
Super-resolution (SR) fluorescence microscopy has been revolutionizing the way in which we investigate the structures, dynamics, and functions of a wide range of nanoscale systems. In this review, I describe the current state of various SR fluorescence microscopy techniques along with the latest developments of fluorophores and labeling for the SR microscopy. I discuss the applications of SR microscopy in the fields of life science and materials science with a special emphasis on quantitative molecular imaging and nanoscale functional imaging. These studies open new opportunities for unraveling the physical, chemical, and optical properties of a wide range of nanoscale architectures together with their nanostructures and will enable the development of new (bio-)nanotechnology.

No MeSH data available.