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Cited by 89Advanced Fluorescence Microscopy TechniquesCellular transport and secretionLipid Membrane Structure and Behavior

Analysis of protein and lipid dynamics using confocal fluorescence recovery after photobleaching (FRAP)

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Day, Charles A., Kraft, Lewis J., Kang, Minchul, Kenworthy, Anne K.·2012·Current Protocols in Cytometry
DOI 10.1002/0471142956.cy0219s62PMC PMC3538152
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Abstract

Fluorescence recovery after photobleaching (FRAP) is a powerful, versatile, and widely accessible tool to monitor molecular dynamics in living cells that can be performed using modern confocal microscopes. Although the basic principles of FRAP are simple, quantitative FRAP analysis requires careful experimental design, data collection, and analysis. In this unit, we discuss the theoretical basis for confocal FRAP, followed by step‐by‐step protocols for FRAP data acquisition using a laser‐scanning confocal microscope for (1) measuring the diffusion of a membrane protein, (2) measuring the diffusion of a soluble protein, and (3) analysis of intracellular trafficking. Finally, data analysis procedures are discussed, and an equation for determining the diffusion coefficient of a molecular species undergoing pure diffusion is presented. Curr. Protoc. Cytom. 62:2.19.1‐2.19.29. © 2012 by John Wiley & Sons, Inc.

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Authors
Day, C. A., Kraft, L. J., Kang, M., & Kenworthy, A. K.
Affiliations
Vanderbilt University (US)
ORCID
Venue
Current Protocols in Cytometry
Publisher
Wiley
Published
2012-10
Volume
62
Issue
1
Pages
Unit2.19-Unit2.19
ISSN
  • 1934-9297 (ISSN-L)
  • 1934-9300
DOI
10.1002/0471142956.cy0219s62
PMC
PMC3538152
OpenAlex
W1592169524
Cited by
89
References
0
Citations by year
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FWCI
1.25
Type
journal article
Language
en
Open access
Licence
onlinelibrary.wiley.com/termsAndConditions#vor (vor, 14 d)
Locations
  • Current Protocols in Cytometry (publishedVersion)
  • PubMed (publishedVersion)
  • Europe PMC (PubMed Central) (acceptedVersion)
  • PubMed Central (acceptedVersion)
Funders
  • American Cancer Society
  • National Institute of General Medical Sciences (T32GM008320, R01GM073846, 3R01 GM73846-4S1)
Topics
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Biophysics › Advanced Fluorescence Microscopy Techniques
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Cell Biology › Cellular transport and secretion
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Lipid Membrane Structure and Behavior
Keywords
FRAP, confocal microscopy, protein diffusion, membrane protein diffusion, intracellular trafficking, diffusion coefficient, lipid dynamics
MeSH terms
Animals, Cell Membrane, Chlorocebus aethiops, Cell Biology, Diffusion, Image Processing, Computer-Assisted, Kinetics, Lipids, Plasmids, Proteins, Software, Models, Statistical, DNA, Complementary, Microscopy, Confocal, COS Cells, Fluorescence Recovery After Photobleaching

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