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J Vis Exp. 2019 Jul 17;(149). doi: 10.3791/59577.

Detecting and Characterizing Protein Self-Assembly In Vivo by Flow Cytometry.

Author information

1
Stowers Institute for Medical Research.
2
Stowers Institute for Medical Research; Department of Molecular and Integrative Physiology, The University of Kansas School of Medicine; rhn@stowers.org.
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Contributed equally

Abstract

Protein self-assembly governs protein function and compartmentalizes cellular processes in space and time. Current methods to study it suffer from low-sensitivity, indirect read-outs, limited throughput, and/or population-level rather than single-cell resolution. We designed a flow cytometry-based single methodology that addresses all of these limitations: Distributed Amphifluoric FRET or DAmFRET. DAmFRET detects and quantifies protein self-assemblies by sensitized emission FRET in vivo, enables deployment across model systems-from yeast to human cells-and achieves sensitive, single-cell, high-throughput read-outs irrespective of protein localization or solubility.

PMID:
31380843
DOI:
10.3791/59577

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