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Single Cell GFP-Trap Reveals Stoichiometry and Dynamics of Cytosolic Protein Complexes

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Department of Biology, University of Osnabrück, 49076 Osnabrück, Germany
Cite this: Nano Lett. 2015, 15, 5, 3610–3615
Publication Date (Web):April 22, 2015
https://doi.org/10.1021/acs.nanolett.5b01153
Copyright © 2015 American Chemical Society

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    Abstract

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    We developed in situ single cell pull-down (SiCPull) of GFP-tagged protein complexes based on micropatterned functionalized surface architectures. Cells cultured on these supports are lysed by mild detergents and protein complexes captured to the surface are probed in situ by total internal reflection fluorescence microscopy. Using SiCPull, we quantitatively mapped the lifetimes of various signal transducer and activator of transcription complexes by monitoring dissociation from the surface and defined their stoichiometry on the single molecule level.

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    Supplementary figures and tables, detailed materials and methods for surface micropatterning, single cell pulldown experiments, and data evaluation. This material is available free of charge via the Internet at http://pubs.acs.org.

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