Measuring ligand-cell surface receptor affinities with axial line-scanning fluorescence correlation spectroscopy release_rev_4d2b6881-681d-44bd-ac81-b53046e3dc8f

by Antonia Franziska Eckert, Peng Gao, Janine Wesslowski, Xianxian Wang, Jasmijn Rath, Karin Nienhaus, Gary Davidson, Gerd Ulrich Nienhaus

Published in eLife by eLife Sciences Publications, Ltd.

2020   Volume 9

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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Type  article-journal
Stage   published
Date   2020-05-22
Language   en ?
DOI  10.7554/elife.55286
PubMed  32441251
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ISSN-L:  2050-084X
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