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  Scanning Fluorescence Correlation Spectroscopy (SFCS) with a Scan Path Perpendicular to the Membrane Plane

Müller, P., Schwille, P., & Weidemann, T. (2014). Scanning Fluorescence Correlation Spectroscopy (SFCS) with a Scan Path Perpendicular to the Membrane Plane. In Y. Engelborghs, & A. Visser (Eds.), Fluorescence Spectroscopy and Microscopy (pp. 635-651). Totowa: Humana Press.

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Genre: Book Chapter
Subtitle : protocol

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 Creators:
Müller, Paul1, Author
Schwille, Petra2, Author              
Weidemann, Thomas2, Author              
Affiliations:
1external, ou_persistent22              
2Schwille, Petra / Cellular and Molecular Biophysics, Max Planck Institute of Biochemistry, Max Planck Society, ou_1565169              

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Free keywords: Scanning fluorescence correlation spectroscopy (SFCS) Fluorescence correlation spectroscopy (FCS) Fluorescence cross-correlation spectroscopy (FCCS) Membrane diffusion Giant unilamellar vesicles (GUV) Diffusion Protein–protein interaction Ligand binding
 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.

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Language(s): eng - English
 Dates: 2014
 Publication Status: Published in print
 Pages: -
 Publishing info: -
 Table of Contents: -
 Rev. Type: -
 Degree: -

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Title: Fluorescence Spectroscopy and Microscopy
  Other : Methods in Molecular Biology (Methods and Protocols), vol 1076
Source Genre: Book
 Creator(s):
Engelborghs, Y., Editor
Visser, A., Editor
Affiliations:
-
Publ. Info: Totowa : Humana Press
Pages: - Volume / Issue: - Sequence Number: - Start / End Page: 635 - 651 Identifier: ISBN: 978-1-62703-648-1

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Title: Methods in Molecular Biology
  Other : Methods in Molecular Biology (Methods and Protocols)
Source Genre: Series
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Publ. Info: Totowa : Humana Press
Pages: - Volume / Issue: 1076 Sequence Number: - Start / End Page: - Identifier: ISSN: 1064-3745