Use of flow cytometric methods to quantify protein-protein interactions

Levi L Blazer; David L Roman; Molly R Muxlow; Richard R Neubig

doi:10.1002/0471142956.cy1311s51

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Use of flow cytometric methods to quantify protein-protein interactions

Journal article

Open access

Use of flow cytometric methods to quantify protein-protein interactions

Levi L Blazer, David L Roman, Molly R Muxlow and Richard R Neubig

Current protocols in cytometry, Vol.Chapter 13(1), pp.Unit 13.11.1-13.11.15

01/2010

DOI: 10.1002/0471142956.cy1311s51

PMCID: PMC2849137

PMID: 20069525

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Abstract

A method is described for the quantitative analysis of protein-protein interactions using the flow cytometry protein interaction assay (FCPIA). This method is based upon immobilizing protein on a polystyrene bead, incubating these beads with a fluorescently labeled binding partner, and assessing the sample for bead-associated fluorescence in a flow cytometer. This method can be used to calculate protein-protein interaction affinities or to perform competition experiments with unlabeled binding partners or small molecules. Examples described in this protocol highlight the use of this assay in the quantification of the affinity of binding partners of the regulator of G-protein signaling protein, RGS19, in either a saturation or a competition format. An adaptation of this method that is compatible for high-throughput screening is also provided.

Flow Cytometry - methods

Biotinylation

High-Throughput Screening Assays

Protein Binding

RGS Proteins - metabolism

Biological Assay

Biotin - metabolism

Details

Title: Subtitle: Use of flow cytometric methods to quantify protein-protein interactions
Creators: Levi L Blazer - Department of Pharmacology, The University of Michigan Medical School, Ann Arbor, Michigan, USA
David L Roman
Molly R Muxlow
Richard R Neubig
Resource Type: Journal article
Publication Details: Current protocols in cytometry, Vol.Chapter 13(1), pp.Unit 13.11.1-13.11.15
DOI: 10.1002/0471142956.cy1311s51
PMID: 20069525
PMCID: PMC2849137
NLM abbreviation: Curr Protoc Cytom
ISSN: 1934-9297
eISSN: 1934-9300
Publisher: United States
Grant note: F32 GM076821 / NIGMS NIH HHS R01 GM039561-20 / NIGMS NIH HHS T32 GM008597 / NIGMS NIH HHS GM076821 / NIGMS NIH HHS F32 GM076821-02 / NIGMS NIH HHS R01 GM039561-21 / NIGMS NIH HHS T32 GM00008597 / NIGMS NIH HHS GM39561 / NIGMS NIH HHS R01 GM039561 / NIGMS NIH HHS T32 GM008597-13A1S1 / NIGMS NIH HHS
Language: English
Date published: 01/2010
Academic Unit: Pharmacy; Iowa Neuroscience Institute; Pharmaceutical Sciences and Experimental Therapeutics; Medicinal and Natural Products Chemistry
Record Identifier: 9984065386302771

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