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Fluorescence Correlation Spectroscopy — Going Beyond the Diffraction Limit

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The combination of short measurement times along with free position or scanning of the observation spot makes fluorescence correlation spectroscopy an excellent tool for investigating diffusion heterogeneity over time and space.

ANDRÉ DEVAUX, ANDREAS BÜLTER and FELIX KOBERLING, PICOQUANT GMBH

A cell membrane is a highly heterogeneous medium containing a wide variety of molecules and structural elements. All of these species may interact with each other, leading to rather complex lateral diffusion behaviors in the lipid bilayer. Depending on the local composition of its membrane neighborhood, a molecule can experience one of four prominent diffusion modes1 (Figure 1). Figure 1. The four principal modes for molecules diffusing in membranes. Molecules are shown as red spots, black lines represent their diffusion paths, and the fluorescence correlation spectroscopy (FCS)...Read full article

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    Published: October 2016
    Glossary
    superresolution
    Superresolution refers to the enhancement or improvement of the spatial resolution beyond the conventional limits imposed by the diffraction of light. In the context of imaging, it is a set of techniques and algorithms that aim to achieve higher resolution images than what is traditionally possible using standard imaging systems. In conventional optical microscopy, the resolution is limited by the diffraction of light, a phenomenon described by Ernst Abbe's diffraction limit. This limit sets a...
    fluorescence correlation spectroscopy
    A powerful method, referred to as FCS, for determining the average diffusion coefficients of fluorescent molecules in solution or membranes. FCS measurements rely on recording the transition of several thousands of molecules through the focal volume. The combination of short measurement times along with free positioning or scanning of the observation spot makes FCS an excellent tool for investigating diffusion heterogeneity over time and space.
    sted microscopy
    STED microscopy, or stimulated emission depletion microscopy, is a super-resolution imaging technique in fluorescence microscopy that surpasses the diffraction limit, enabling the visualization of structures at the nanoscale level. This technique was developed to overcome the limitations imposed by the diffraction of light, which traditionally hindered the resolution of optical microscopy to a few hundred nanometers. Key features and principles of STED microscopy: Super-resolution: STED...
    diode lasersspectroscopysuperresolutionMicroscopyLasersEuropeBiophotonicsfluorescence correlation spectroscopyFCSstimulated emission depletion microscopySTED microscopyconfocal microscopydiffusion coefficientmolecular biologydiffraction limitPicoQuantAndré DevauxAndreas BülterFelix Koberlingtime-correlatedsingle-photon countingTCSPCpulsed interleaved excitationPiEFeatures

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