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Cell-Surface Protein–Protein Interaction Analysis with Time-Resolved FRET and Snap-Tag Technologies

  • Timothy N. Feinstein
Protocol
  • 2.8k Downloads
Part of the Methods in Molecular Biology book series (MIMB, volume 1066)

Abstract

Förster resonance energy transfer (FRET) is a proximity-dependent quantum effect that allows the measurement of protein interactions and conformational changes which are invisible to traditional forms of fluorescence or electron microscopy. However, FRET experiments often have difficulty detecting interactions that are transient and localized or occur in low abundance against a large background. This protocol describes a method of improving on the sensitivity and quantifiability of FRET experiments by using time-specific detection to isolate FRET-mediated acceptor emission from cross-talk excitation and all other sources of nonspecific fluorescence background.

Key words

Förster resonance energy transfer FRET Protein–protein interactions SNAP tagging G-protein-coupled receptors GPCR 

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Copyright information

© Springer Science+Business Media, New York 2013

Authors and Affiliations

  • Timothy N. Feinstein
    • 1
  1. 1.Department of Pharmacology and Chemical BiologyUniversity of PittsburghPittsburghUSA

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