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Related Concept Videos

Quantification of Protein Interaction Network Dynamics using Multiplexed Co-Immunoprecipitation07:57

Quantification of Protein Interaction Network Dynamics using Multiplexed Co-Immunoprecipitation

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Quantitative Multiplex Immunoprecipitation (QMI) uses flow cytometry for sensitive detection of differences in the abundance of targeted protein-protein interactions between two samples. QMI can be performed using a small amount of biomaterial, does not require genetically engineered tags, and can be adapted for any previously defined protein interaction...
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Protein Networks02:26

Protein Networks

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An organism can have thousands of different proteins, and these proteins must cooperate to ensure the health of an organism. Proteins bind to other proteins and form complexes to carry out their functions. Many proteins interact with multiple other proteins creating a complex network of protein interactions.
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This video demonstrates co-immunoprecipitated protein complexes on a PVDF membrane for protein-protein interaction analysis. The reduced proteins from complexes were treated with trypsin to cleave the individual proteins at the smaller peptides, then, the remaining peptides were extracted from the PVDF membrane. The pulled peptide derived from both proteins in the complex was then dried and resuspended in a low concentration of formic acid for further...
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Protein-protein interactions are important for elucidating the function of target proteins, and co-immunoprecipitation (co-IP) can easily confirm PPIs. We transiently transfected a plasmid encoding an epitope-tagged protein into HEK-293 cells and developed an immunoprecipitation method to easily confirm the binding of two target...
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Related Experiment Video

Updated: Jan 19, 2026

Quantification of Protein Interaction Network Dynamics using Multiplexed Co-Immunoprecipitation
07:57

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Quantification of Protein Interaction Network Dynamics using Multiplexed Co-Immunoprecipitation.

Emily A Brown1, Steven C Neier2, Claudia Neuhauser3

  • 1Center for Integrative Brain Research, Seattle Children's Research Institute; Graduate Program in Neuroscience, University of Washington.

Journal of Visualized Experiments : Jove
|September 10, 2019
PubMed
Summary

Quantitative Multiplex Immunoprecipitation (QMI) offers a novel method to measure dynamic protein interactions. This technique quantifies protein complexes using flow cytometry, enabling new insights into cellular signaling networks.

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Area of Science:

  • Cellular Biology
  • Molecular Interactions
  • Biochemistry

Background:

  • Dynamic protein-protein interactions are crucial for cellular functions, but their study in complex networks is challenging.
  • Existing methods for monitoring protein interactions often require significant biomaterial or genetic manipulation.

Purpose of the Study:

  • To present a protocol for Quantitative Multiplex Immunoprecipitation (QMI) for the quantitative assessment of protein-protein interactions.
  • To enable the study of dynamic changes in protein interaction networks with high sensitivity and minimal sample input.

Main Methods:

  • Quantitative Multiplex Immunoprecipitation (QMI) uses flow cytometry to measure Proteins in Shared Complexes detected by Exposed Surface epitopes (PiSCES).
  • Immunoprecipitation antibodies are conjugated to spectrally distinct MagBeads, allowing multiplexed detection of protein complexes.
  • Probe antibodies are labeled with fluorophores to quantify protein interaction abundance.

Main Results:

  • QMI allows quantitative assessment of fold changes in protein interactions without genetic tagging.
  • The method requires minimal biomaterial and can differentiate multiple parallel immunoprecipitations simultaneously.
  • QMI has been successfully applied to T cell signaling and neuronal synapses, generating hypotheses for diagnostic and therapeutic applications.

Conclusions:

  • QMI provides a robust and versatile protocol for studying dynamic protein-protein interactions.
  • This method facilitates the characterization of complex signaling networks and has potential for biomarker discovery.
  • The protocol is adaptable for various protein interaction studies, offering a valuable tool for molecular biology research.