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Author Notes:

vfaunde@emory.edu; gbassel@emory.edu

A.N.V. and A.G. contributed to the writing of the manuscript; V.F. and G.J.B. edited the manuscript. Figures generated by A.N.V. Initial optimization by A.G. Lab infrastructure and resources by G.J.B. and V.F.

The authors declare no competing interests.

Subjects:

Research Funding:

This work was funded by NIH R01 MH109026 (G.J.B.), F31 NS101932 (A.N.V.), 1R56MH111459 (V.F.), and 1RF1AG060285 (V.F.).

Protocol for Immuno-Enrichment of FLAG-Tagged Protein Complexes

Tools:

Journal Title:

STAR Protocols

Volume:

Volume 1, Number 2

Publisher:

Type of Work:

Article | Final Publisher PDF

Abstract:

This protocol describes immunoprecipitation of proteins associated with FLAG-tagged recombinant proteins followed by mass spectrometry-based proteomics to identify the associated interactome components. FLAG epitope was chosen, because existing high-affinity monoclonal antibodies allow for sensitive immunoprecipitation and FLAG peptides permit efficient elution of protein complexes. With many commercially available FLAG tools, this protocol is highly versatile. This procedure reduces immunoprecipitation of nonspecific binding proteins. Gene ontology analyses performed following mass spectrometry-based proteomics may elucidate novel functions of proteins of interest.

Copyright information:

© 2020 The Authors

This is an Open Access work distributed under the terms of the Creative Commons Attribution-NonCommercial-NoDerivatives 4.0 International License (https://creativecommons.org/licenses/by-nc-nd/4.0/).
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