Imaging Protein‐Protein Interactions by Förster Resonance Energy Transfer (FRET) Microscopy in Live Cells. (9th July 2018)

• Record Type: Journal Article
• Authors: Manzella‐Lapeira, Javier; Brzostowski, Joseph A.
• Editors: Coligan, John E.; Dunn, Ben M.; Speicher, David W.; Wingfield, Paul T.
• Is Part Of: Current protocols in protein science. Volume 93(2018)
• Page Start: n/a
• Page End: n/a
• Publication Date: 2018-07-09
• Subjects: confocal microscopy -- FRET -- FLIM -- live cell imaging; Proteins -- Laboratory manuals; Proteins; Clinical Laboratory Techniques; Genetic Techniques; Immunologic Techniques; Proteins; Laboratory manuals; 572.6028
• DOI: 10.1002/cpps.58
• Languages: English
• ISSNs: 1934-3655

Abstract

Abstract: This updated unit compares three methods to acquire Förster Resonance Energy Transfer (FRET) data in living cells using a confocal microscope: Acceptor photobleaching, Acceptor‐sensitized emission FRET, and Donor fluorescence lifetime imaging. Detailed protocols for live cell husbandry, image acquisition, and data analysis are provided. In addition to providing instructions for manufacturer's analysis tool sets, we provide an easy‐to‐use, MATLAB‐based code to calculate FRET efficiency from data obtained using the Acceptor photobleaching or Acceptor‐sensitized emission method, which can be freely downloaded. © 2018 by John Wiley & Sons, Inc.