1. Physics of Living Systems
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Measuring ligand-cell surface receptor affinities with axial line-scanning fluorescence correlation spectroscopy

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Cite this article as: eLife 2020;9:e55286 doi: 10.7554/eLife.55286

Abstract

Development and homeostasis of multicellular organisms is largely controlled by complex cell-cell signaling networks that rely on specific binding of secreted ligands to cell surface receptors. The Wnt signaling network, as an example, involves multiple ligands and receptors to elicit specific cellular responses. To understand the mechanisms of such a network, ligand-receptor interactions should be characterized quantitatively, ideally in live cells or tissues. Such measurements are possible using fluorescence microscopy yet challenging due to sample movement, low signal-to-background ratio and photobleaching. Here we present a robust approach based on fluorescence correlation spectroscopy with ultra-high speed axial line scanning, yielding precise equilibrium dissociation coefficients of interactions in the Wnt signaling pathway. Using CRISPR/Cas9 editing to endogenously tag receptors with fluorescent proteins, we demonstrate that the method delivers precise results even with low, near-native amounts of receptors.

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Author details

  1. Antonia Franziska Eckert

    Institute of Applied Physics, Karlsruhe Institute of Technology, Karlsruhe, Germany
    Competing interests
    The authors declare that no competing interests exist.
  2. Peng Gao

    Institute of Applied Physics, Karlsruhe Institute of Technology, Karlsruhe, Germany
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-5354-3944
  3. Janine Wesslowski

    Institute of Biological and Chemical Systems, Karlsruhe Institute of Technology, Karlsruhe, Germany
    Competing interests
    The authors declare that no competing interests exist.
  4. Xianxian Wang

    Institute of Biological and Chemical Systems, Karlsruhe Institute of Technology, Karlsruhe, Germany
    Competing interests
    The authors declare that no competing interests exist.
  5. Jasmijn Rath

    Institute of Applied Physics, Karlsruhe Institute of Technology, Karlsruhe, Germany
    Competing interests
    The authors declare that no competing interests exist.
  6. Karin Nienhaus

    Institute of Applied Physics, Karlsruhe Institute of Technology, Karlsruhe, Germany
    Competing interests
    The authors declare that no competing interests exist.
  7. Gary Davidson

    Institute of Biological and Chemical Systems, Karlsruhe Institute of Technology, Karlsruhe, Germany
    For correspondence
    gary.davidson@kit.edu
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-2264-5518
  8. Gerd Ulrich Nienhaus

    Institute of Applied Physics, Karlsruhe Institute of Technology, Karlsruhe, Germany
    For correspondence
    uli@uiuc.edu
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-5027-3192

Funding

Deutsche Forschungsgemeinschaft (SFB 1324,project A6,project Z2)

  • Gerd Ulrich Nienhaus

Deutsche Forschungsgemeinschaft (SFB 1324,project A6)

  • Gary Davidson

Helmholtz-Gemeinschaft (STN)

  • Gerd Ulrich Nienhaus

Helmholtz-Gemeinschaft (BIFTM)

  • Gary Davidson

The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.

Reviewing Editor

  1. Hannes Neuweiler, University of Würzburg, Germany

Publication history

  1. Received: January 20, 2020
  2. Accepted: May 21, 2020
  3. Accepted Manuscript published: May 22, 2020 (version 1)

Copyright

© 2020, Eckert et al.

This article is distributed under the terms of the Creative Commons Attribution License permitting unrestricted use and redistribution provided that the original author and source are credited.

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