The Experts below are selected from a list of 18945 Experts worldwide ranked by ideXlab platform

Masataka Kinjo - One of the best experts on this subject based on the ideXlab platform.

  • rotational diffusion measurements using polarization dependent Fluorescence Correlation Spectroscopy based on superconducting nanowire single photon detector
    Optics Express, 2015
    Co-Authors: Johtaro Yamamoto, Taro Yamashita, Shigehito Miki, Tokuko Haraguchi, Yasushi Hiraoka, Hirotaka Terai, Makoto Oura, Takashi Jin, Masataka Kinjo
    Abstract:

    Conventional polarization-dependent Fluorescence Correlation Spectroscopy (pol-FCS) requires two sets of photon detectors to eliminate after-pulse noises (dual-channel pol-FCS; DC-pol-FCS) in the sub-microsecond range. In this study, we successfully realized pol-FCS using a visible-wavelength superconductive nanowire single-photon detector (single-channel pol-FCS; SC-pol-FCS). The detector used is free of after-pulse noises and thus eliminates the need for dual channels in pol-FCS. Further, the optics in the SC-pol-FCS system are easier to adjust than those in the conventional system. Consequently, we obtained higher signal-to-noise ratios compared with conventional DC-pol-FCS systems. Thus, SC-pol-FCS is a potentially useful system for obtaining pol-FCS measurements, and can facilitate improved rotational diffusion studies.

  • Fluorescence Correlation Spectroscopy with visible wavelength superconducting nanowire single photon detector
    2015 15th International Superconductive Electronics Conference (ISEC), 2015
    Co-Authors: Taro Yamashita, Masataka Kinjo, Shigehito Miki, Johtaro Yamamoto, Tokuko Haraguchi, Hirotaka Terai, Yasushi Hiraoka
    Abstract:

    We present the Fluorescence Correlation Spectroscopy (FCS) using superconducting nanowire single-photon detectors (SSPDs) which is after-pulse free unlike the avalanche photodiode (APD). Multimode-fiber-coupled SSPDs with high system detection efficiency for visible wavelengths were developed and implemented in the FCS system. We performed FCS measurements for Rhodamine 6G as fluorescent samples, and demonstrated that autoCorrelation functions obtained by the SSPDs showed an after-pulse-free curve in the short Correlation time region of sub microseconds where the afterpulse effect was dominant using the APD. Also for the longer Correlation time region over microseconds, the SSPD revealed the triplet component of Rhodamine 6G, which is obscured by the APD due to after-pulse effect. The obtained results clearly indicate the advantage of SSPDs for the FCS system.

  • Fluorescence Correlation Spectroscopy with visible wavelength superconducting nanowire single photon detector
    Optics Express, 2014
    Co-Authors: Taro Yamashita, Masataka Kinjo, Dengkuan Liu, Shigehito Miki, Johtaro Yamamoto, Tokuko Haraguchi, Yasushi Hiraoka, Zhen Wang, Hirotaka Terai
    Abstract:

    We present the first demonstration of Fluorescence Correlation Spectroscopy (FCS) using superconducting nanowire single-photon detectors (SSPDs) which are free of afterpulses unlike the avalanche photodiode (APD). Multimode-fiber-coupled SSPDs with high system detection efficiency for visible wavelengths were developed and implemented in the FCS system. We performed FCS measurements for Rhodamine B and 6G as fluorescent samples, and found that autoCorrelation functions obtained by the SSPDs showed a noise-free curve in the short Correlation time region of sub microseconds where the afterpulse effect was dominant using the APD. The obtained results clearly indicate the advantage of SSPDs for the FCS system.

  • multipoint Fluorescence Correlation Spectroscopy with total internal reflection Fluorescence microscope
    Journal of Biomedical Optics, 2009
    Co-Authors: Yu Ohsugi, Masataka Kinjo
    Abstract:

    We report simultaneous determination of diffusion coefficients at different points of a cell membrane using a multipoint Fluorescence Correlation Spectroscopy (FCS) system. A system carrying seven detection areas in the evanescent field is achieved by using seven optical fibers on the image plane in the detection port of an objective-type total internal reflection FCS (TIR-FCS) system. Fluctuation of Fluorescence intensity is monitored and evaluated using seven photomultiplier tubes (PMTs) and a newly constructed multichannel correlator. We demonstrate simultaneous-multipoint FCS, with a 3-µs time resolution, to investigate heterogeneous structures such as cell membranes and membrane-binding molecular dynamics near glass surfaces in live cells.

  • analysis of membrane binding protein mobility in living cells using total internal reflection Fluorescence Correlation Spectroscopy
    Biophysical Reviews and Letters, 2006
    Co-Authors: Yu Ohsugi, Masataka Kinjo
    Abstract:

    Total internal reflection Fluorescence Correlation Spectroscopy (TIR-FCS) is an appropriate method for measuring diffusion constants and the number of fluorescent molecules very close to the coverglass surface. Recently, we have reported the application of TIR-FCS to cell biology, measuring membrane-binding farnesylated green fluorescent proteins (EGFP-F) in living cells. In this research, we measured the signal transduction molecule, protein kinase C (PKC), fused with EGFP in living HeLa cells by using TIR-FCS. We observed two different diffusional mobilities of PKCβII-EGFP, three-dimensional faster diffusion near the plasma membrane and slower lateral diffusion on the plasma membrane after adinosine tri phosphate (ATP) activation. These results indicate that it is possible to use TIR-FCS in the study of molecular dynamics and interactions of signal transduction proteins on the plasma membrane of the living cell.

Theo Lasser - One of the best experts on this subject based on the ideXlab platform.

  • high volume confinement in two photon total internal reflection Fluorescence Correlation Spectroscopy
    Applied Physics Letters, 2009
    Co-Authors: Denis A Ivanov, Marcel Dr Leutenegger, Vladislav I Shcheslavskiy, Iwan Marki, Theo Lasser
    Abstract:

    We report results on two-photon total-internal-reflection Fluorescence Correlation Spectroscopy with radially polarized light. The combination of liquid crystal spatial light modulator, providing radial polarization with ultrafast laser (picosecond Nd:GdVO4 laser), allowed us to take the advantage of nonlinear optical contrast mechanisms to suppress the side-lobe energy specific for radial polarization and reduce the effective excited volume twice compared to one-photon evanescent wave excitation in Fluorescence Correlation Spectroscopy.

  • high volume confinement in two photon Fluorescence Correlation Spectroscopy with radially polarized light
    Bios, 2009
    Co-Authors: Denis A Ivanov, Marcel Dr Leutenegger, Vladislav I Shcheslavskiy, Iwan Marki, Theo Lasser
    Abstract:

    We present the results on two-photon total-internal-reflection Fluorescence Correlation Spectroscopy. The combination of liquid crystal spatial light modulator, providing radial polarization, with ultrafast laser (picosecond Nd:GdVO4 laser) allowed us to take the advantage of nonlinear optical contrast mechanisms to suppress the side-lobe energy specific for radial polarization and reduce the effective excited volume twice compared to one-photon evanescent wave excitation in Fluorescence Correlation Spectroscopy.

  • dynamic disorder in horseradish peroxidase observed with total internal reflection Fluorescence Correlation Spectroscopy
    Optics Express, 2007
    Co-Authors: Kai Hassler, Rudolf Rigler, Hans Blom, Jerker Widengren, Per Rigler, Theo Lasser
    Abstract:

    This paper discusses the application of objective-type total internal reflection Fluorescence Correlation Spectroscopy (TIR-FCS) to the study of the kinetics of immobilized horseradish peroxidase on a single molecule level. Objective-type TIR-FCS combines the advantages of FCS with TIRF microscopy in a way that allows for simultaneous ultra-sensitive spectroscopic measurements using a single-point detector and convenient localization of single molecules on a surface by means of parallel imaging.

  • confining the sampling volume for Fluorescence Correlation Spectroscopy using a sub wavelength sized aperture
    Optics Express, 2006
    Co-Authors: Marcel Dr Leutenegger, Alexandre Perentes, Michael Gösch, Olivier J. F. Martin, Patrik Hoffmann, Theo Lasser
    Abstract:

    For the observation of single molecule dynamics with Fluorescence fluctuation Spectroscopy (FFS) very low fluorophore concentrations are necessary. For in vitro measurements, this requirement is easy to fulfill. In biology however, micromolar concentrations are often encountered and may pose a real challenge to conventional FFS methods based on confocal instrumentation. We show a higher confinement of the sampling volume in the near-field of sub-wavelength sized apertures in a thin gold film. The gold apertures have been measured and characterized with Fluorescence Correlation Spectroscopy (FCS), indicating light confinement beyond the far-field diffraction limit. We measured a reduction of the effective sampling volume by an order of magnitude compared to confocal instrumentation.

  • fcs Fluorescence Correlation Spectroscopy tir fcs total internal reflection fcs
    2006
    Co-Authors: Matthias Geissbuhler, Marcel Dr Leutenegger, Christian Eggeling, Iwan Maerki, Hans Blom, Theo Lasser
    Abstract:

    Abstract We present the development and application of a dual-color setup that can be used for confocal Fluorescence Correlation Spectroscopy (FCS), total internal reflection FCS (TIR-FCS) as well as dual color imaging.As exemplary application we performed preliminary measure-ments for DNA-Sequencing with a novel surface chemistry. Motivation • Single molecule detection of chemical kinetics A(red)+B(blue) AB• Monitoring biological processes (eg molecular binding essays)• in vivo observations of dynamics of molecular motors Application functionalized surfaceLinker to surfacecircularized singleDNA strandconfined excitation anddetection Volumecompleted DNA strandEnzymefree Nucleotides DNA Sequencing • improved surface chemistry• bind single DNA strand to surface• labeling nucleotides• observation in confocal volume Figure 4: DNA Sequencing methodFigure 5: Image of dual colored spots afterseveral minutes of nucleotide-incorporation Outlook • Advanced concepts for further confined sampling volumes• Higher concentrations of labeled molecules• More colors or/and lifetime seperation• More biological systems

Hirotaka Terai - One of the best experts on this subject based on the ideXlab platform.

  • rotational diffusion measurements using polarization dependent Fluorescence Correlation Spectroscopy based on superconducting nanowire single photon detector
    Optics Express, 2015
    Co-Authors: Johtaro Yamamoto, Taro Yamashita, Shigehito Miki, Tokuko Haraguchi, Yasushi Hiraoka, Hirotaka Terai, Makoto Oura, Takashi Jin, Masataka Kinjo
    Abstract:

    Conventional polarization-dependent Fluorescence Correlation Spectroscopy (pol-FCS) requires two sets of photon detectors to eliminate after-pulse noises (dual-channel pol-FCS; DC-pol-FCS) in the sub-microsecond range. In this study, we successfully realized pol-FCS using a visible-wavelength superconductive nanowire single-photon detector (single-channel pol-FCS; SC-pol-FCS). The detector used is free of after-pulse noises and thus eliminates the need for dual channels in pol-FCS. Further, the optics in the SC-pol-FCS system are easier to adjust than those in the conventional system. Consequently, we obtained higher signal-to-noise ratios compared with conventional DC-pol-FCS systems. Thus, SC-pol-FCS is a potentially useful system for obtaining pol-FCS measurements, and can facilitate improved rotational diffusion studies.

  • Fluorescence Correlation Spectroscopy with visible wavelength superconducting nanowire single photon detector
    2015 15th International Superconductive Electronics Conference (ISEC), 2015
    Co-Authors: Taro Yamashita, Masataka Kinjo, Shigehito Miki, Johtaro Yamamoto, Tokuko Haraguchi, Hirotaka Terai, Yasushi Hiraoka
    Abstract:

    We present the Fluorescence Correlation Spectroscopy (FCS) using superconducting nanowire single-photon detectors (SSPDs) which is after-pulse free unlike the avalanche photodiode (APD). Multimode-fiber-coupled SSPDs with high system detection efficiency for visible wavelengths were developed and implemented in the FCS system. We performed FCS measurements for Rhodamine 6G as fluorescent samples, and demonstrated that autoCorrelation functions obtained by the SSPDs showed an after-pulse-free curve in the short Correlation time region of sub microseconds where the afterpulse effect was dominant using the APD. Also for the longer Correlation time region over microseconds, the SSPD revealed the triplet component of Rhodamine 6G, which is obscured by the APD due to after-pulse effect. The obtained results clearly indicate the advantage of SSPDs for the FCS system.

  • Fluorescence Correlation Spectroscopy with visible wavelength superconducting nanowire single photon detector
    Optics Express, 2014
    Co-Authors: Taro Yamashita, Masataka Kinjo, Dengkuan Liu, Shigehito Miki, Johtaro Yamamoto, Tokuko Haraguchi, Yasushi Hiraoka, Zhen Wang, Hirotaka Terai
    Abstract:

    We present the first demonstration of Fluorescence Correlation Spectroscopy (FCS) using superconducting nanowire single-photon detectors (SSPDs) which are free of afterpulses unlike the avalanche photodiode (APD). Multimode-fiber-coupled SSPDs with high system detection efficiency for visible wavelengths were developed and implemented in the FCS system. We performed FCS measurements for Rhodamine B and 6G as fluorescent samples, and found that autoCorrelation functions obtained by the SSPDs showed a noise-free curve in the short Correlation time region of sub microseconds where the afterpulse effect was dominant using the APD. The obtained results clearly indicate the advantage of SSPDs for the FCS system.

Petra Schwille - One of the best experts on this subject based on the ideXlab platform.

  • scanning Fluorescence Correlation Spectroscopy sfcs with a scan path perpendicular to the membrane plane
    arXiv: Biomolecules, 2018
    Co-Authors: Paul Muller, Petra Schwille, Thomas Weidemann
    Abstract:

    Scanning Fluorescence Correlation Spectroscopy (SFCS) with a scan path perpendicular to the membrane plane was introduced to measure diffusion and interactions of fluorescent components in free standing biomembranes. Using a confocal laser scanning microscope (CLSM) the open detection volume is moved laterally with kHz frequency through the membrane and the photon events are continuously recorded and stored in a file. While the accessory hardware requirements for a conventional CLSM are minimal, data evaluation can pose a bottleneck. The photon events must be assigned to each scan, in which the maximum signal intensities have to be detected, binned, and aligned between the scans, in order to derive the membrane related intensity fluctuations of one spot. Finally, this time-dependent signal must be correlated and evaluated by well known FCS model functions. Here we provide two platform independent, open source software tools (PyScanFCS and PyCorrFit) that allow to perform all of these steps and to establish perpendicular SFCS in its one- or two-focus as well as its single- or dual-colour modality.

  • scanning Fluorescence Correlation Spectroscopy sfcs with a scan path perpendicular to the membrane plane
    Methods of Molecular Biology, 2014
    Co-Authors: Paul Muller, Petra Schwille, Thomas Weidemann
    Abstract:

    Scanning Fluorescence Correlation Spectroscopy (SFCS) with a scan path perpendicular to the membrane plane was introduced to measure diffusion and interactions of fluorescent components in free-standing biomembranes. Using a confocal laser scanning microscope (CLSM), the open detection volume is repeatedly scanned through the membrane at a kHz frequency. The Fluorescence photons emitted from the detection volume are continuously recorded and stored in a file. While the accessory hardware requirements for a conventional CLSM are minimal, data evaluation can pose a bottleneck. The photon events must be assigned to each scan, in which the maximum signal intensities have to be detected, binned, and aligned between the scans, in order to derive the membrane-related intensity fluctuations of one spot. Finally, this time-dependent signal must be correlated and evaluated by well-known FCS model functions. Here we provide two platform-independent, open source software tools (PyScanFCS and PyCorrFit) that allow to perform all of these steps and to establish perpendicular SFCS in its one- or two-focus as well as its single- or dual-color modality.

  • total internal reflection Fluorescence Correlation Spectroscopy effects of lateral diffusion and surface generated Fluorescence
    Biophysical Journal, 2008
    Co-Authors: Jonas Ries, Eugene P Petrov, Petra Schwille
    Abstract:

    Fluorescence Correlation Spectroscopy with total internal reflection excitation (TIR-FCS) is a promising method with emerging biological applications for measuring binding dynamics of fluorescent molecules to a planar substrate as well as diffusion coefficients and concentrations at the interface. Models for Correlation functions proposed so far are rather approximate for most conditions, since they neglect lateral diffusion of fluorophores. Here we propose accurate extensions of previously published models for axial Correlation functions taking into account lateral diffusion through detection profiles realized in typical experiments. In addition, we consider the effects of surface-generated emission in objective-based TIR-FCS. The expressions for Correlation functions presented here will facilitate quantitative and accurate measurements with TIR-FCS.

  • precise measurement of diffusion coefficients using scanning Fluorescence Correlation Spectroscopy
    Biophysical Journal, 2008
    Co-Authors: Zdeněk Petrasek, Petra Schwille
    Abstract:

    We have implemented scanning Fluorescence Correlation Spectroscopy (sFCS) for precise determination of diffusion coefficients of fluorescent molecules in solution. The measurement volume where the molecules are excited, and from which the Fluorescence is detected, was scanned in a circle with radius comparable to its size at frequencies 0.5–2 kHz. The scan radius R, determined with high accuracy by careful calibration, provides the spatial measure required for the determination of the diffusion coefficient D, without the need to know the exact size of the measurement volume. The difficulties in the determination of the measurement volume size have limited the application of standard FCS with fixed measurement volume to relative measurements, where the diffusion coefficient is determined by comparison with a standard. We demonstrate, on examples of several common fluorescent dyes, that sFCS can be used to measure D with high precision without a need for a standard. The correct value of D can be determined in the presence of weak photobleaching, and when the measurement volume size is modified, indicating the robustness of the method. The applicability of the presented implementation of sFCS to biological systems in demonstrated on the measurement of the diffusion coefficient of eGFP in the cytoplasm of HeLa cells. With the help of simulations, we find the optimal value of the scan radius R for the experiment.

  • Fluorescence Correlation Spectroscopy novel variations of an established technique
    Annual Review of Biophysics and Biomolecular Structure, 2007
    Co-Authors: Elke Haustein, Petra Schwille
    Abstract:

    Fluorescence Correlation Spectroscopy (FCS) is one of the major biophysical techniques used for unraveling molecular interactions in vitro and in vivo. It allows minimally invasive study of dynamic processes in biological specimens with extremely high temporal and spatial resolution. By recording and correlating the Fluorescence fluctuations of single labeled molecules through the exciting laser beam, FCS gives information on molecular mobility and photophysical and photochemical reactions. By using dual-color Fluorescence cross-Correlation, highly specific binding studies can be performed. These have been extended to four reaction partners accessible by multicolor applications. Alternative detection schemes shift accessible time frames to slower processes (e.g., scanning FCS) or higher concentrations (e.g., TIR-FCS). Despite its long tradition, FCS is by no means dated. Rather, it has proven to be a highly versatile technique that can easily be adapted to solve specific biological questions, and it continues to find exciting applications in biology and medicine.

Taro Yamashita - One of the best experts on this subject based on the ideXlab platform.

  • rotational diffusion measurements using polarization dependent Fluorescence Correlation Spectroscopy based on superconducting nanowire single photon detector
    Optics Express, 2015
    Co-Authors: Johtaro Yamamoto, Taro Yamashita, Shigehito Miki, Tokuko Haraguchi, Yasushi Hiraoka, Hirotaka Terai, Makoto Oura, Takashi Jin, Masataka Kinjo
    Abstract:

    Conventional polarization-dependent Fluorescence Correlation Spectroscopy (pol-FCS) requires two sets of photon detectors to eliminate after-pulse noises (dual-channel pol-FCS; DC-pol-FCS) in the sub-microsecond range. In this study, we successfully realized pol-FCS using a visible-wavelength superconductive nanowire single-photon detector (single-channel pol-FCS; SC-pol-FCS). The detector used is free of after-pulse noises and thus eliminates the need for dual channels in pol-FCS. Further, the optics in the SC-pol-FCS system are easier to adjust than those in the conventional system. Consequently, we obtained higher signal-to-noise ratios compared with conventional DC-pol-FCS systems. Thus, SC-pol-FCS is a potentially useful system for obtaining pol-FCS measurements, and can facilitate improved rotational diffusion studies.

  • Fluorescence Correlation Spectroscopy with visible wavelength superconducting nanowire single photon detector
    2015 15th International Superconductive Electronics Conference (ISEC), 2015
    Co-Authors: Taro Yamashita, Masataka Kinjo, Shigehito Miki, Johtaro Yamamoto, Tokuko Haraguchi, Hirotaka Terai, Yasushi Hiraoka
    Abstract:

    We present the Fluorescence Correlation Spectroscopy (FCS) using superconducting nanowire single-photon detectors (SSPDs) which is after-pulse free unlike the avalanche photodiode (APD). Multimode-fiber-coupled SSPDs with high system detection efficiency for visible wavelengths were developed and implemented in the FCS system. We performed FCS measurements for Rhodamine 6G as fluorescent samples, and demonstrated that autoCorrelation functions obtained by the SSPDs showed an after-pulse-free curve in the short Correlation time region of sub microseconds where the afterpulse effect was dominant using the APD. Also for the longer Correlation time region over microseconds, the SSPD revealed the triplet component of Rhodamine 6G, which is obscured by the APD due to after-pulse effect. The obtained results clearly indicate the advantage of SSPDs for the FCS system.

  • Fluorescence Correlation Spectroscopy with visible wavelength superconducting nanowire single photon detector
    Optics Express, 2014
    Co-Authors: Taro Yamashita, Masataka Kinjo, Dengkuan Liu, Shigehito Miki, Johtaro Yamamoto, Tokuko Haraguchi, Yasushi Hiraoka, Zhen Wang, Hirotaka Terai
    Abstract:

    We present the first demonstration of Fluorescence Correlation Spectroscopy (FCS) using superconducting nanowire single-photon detectors (SSPDs) which are free of afterpulses unlike the avalanche photodiode (APD). Multimode-fiber-coupled SSPDs with high system detection efficiency for visible wavelengths were developed and implemented in the FCS system. We performed FCS measurements for Rhodamine B and 6G as fluorescent samples, and found that autoCorrelation functions obtained by the SSPDs showed a noise-free curve in the short Correlation time region of sub microseconds where the afterpulse effect was dominant using the APD. The obtained results clearly indicate the advantage of SSPDs for the FCS system.