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Determination of Lipid Raft Partitioning of Fluorescently-tagged Probes in Living Cells by Fluorescence Correlation Spectroscopy (FCS)
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JoVE Journal Biology
Determination of Lipid Raft Partitioning of Fluorescently-tagged Probes in Living Cells by Fluorescence Correlation Spectroscopy (FCS)

Determination of Lipid Raft Partitioning of Fluorescently-tagged Probes in Living Cells by Fluorescence Correlation Spectroscopy (FCS)

DOI:
10.3791/3513-v

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10:59 min

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April 06, 2012

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, ,
1Centre de Recherche de l’Institut du Cerveau et de la Moelle Épinière,Hôpital de la Pitié-Salpêtrière, 2Institut des Sciences Moléculaires d’Orsay,Université Paris-Sud, 3Centre de Photonique Biomédicale du Centre Laser,Université Paris-Sud

Chapters

  • 00:05Title
  • 01:48Calibration of the FCS Setup
  • 03:45FCS Data Analysis
  • 05:50Staining of Living Cells with Lipid-rafts Marker
  • 06:51FCS Data Acquisition on Living Cells
  • 08:07Representative FCS Analysis
  • 10:18Conclusion

Summary

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A technique to probe the lipid raft partitioning of fluorescent proteins at the plasma membrane of living cells is described. It takes advantage of the disparity in diffusion times of proteins located inside or outside of lipid rafts. Acquisition can be performed dynamically in control conditions or after drug addition.

Tags

Lipid Raft PartitioningFluorescently-tagged ProbesLiving CellsFluorescence Correlation SpectroscopyCell MembranesMicrodomainsCholesterolSphingolipidsSignallingTraffickingDiseasesControversyLipid Raft SizeMicroscopy Resolution LimitDirect ImagingTechniquesDetergent Resistant MembranesCo-patching With AntibodiesArtefactsSensitive AnalysisPlasma Membrane

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