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Single-Molecule Imaging of Lateral Mobility and Ion Channel Activity in Lipid Bilayers using Total Internal Reflection Fluorescence (TIRF) Microscopy
Journal JoVE
Biochimie
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Journal JoVE Biochimie
Single-Molecule Imaging of Lateral Mobility and Ion Channel Activity in Lipid Bilayers using Total Internal Reflection Fluorescence (TIRF) Microscopy

Single-Molecule Imaging of Lateral Mobility and Ion Channel Activity in Lipid Bilayers using Total Internal Reflection Fluorescence (TIRF) Microscopy

DOI:
10.3791/64970-v

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08:55 min

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February 17, 2023

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,
1Biophysics Department, Institute of Biomaterials and Biomolecular Systems,University of Stuttgart

Chapitres

  • 00:05Introduction
  • 00:43Preparation of Lipids and Agarose Hydrogel
  • 01:33PMMA Chamber Assembly and Lipid Monolayer Formation
  • 03:06Preparation of Lipid-Coated Aqueous Droplets in Hexadecane/Silicone Oil Solution
  • 04:17Preparation and Imaging of Single Ion Channels in DIB Membranes
  • 06:49Results: Isolation of TOM-CC and Correlation of Channel Activity with Lateral Mobility of TOM-CC
  • 08:18Conclusion

Summary

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This protocol describes how to use TIRF microscopy to track individual ion channels and determine their activity in supported lipid membranes, thereby defining the interplay between lateral membrane movement and channel function. It describes how to prepare the membranes, record the data, and analyze the results.

Tags

Keywords: Single-molecule ImagingLateral MobilityIon Channel ActivityLipid BilayersTotal Internal Reflection Fluorescence (TIRF) MicroscopyMembrane ProteinsDPhPCHexadecaneSilicone OilAgarose HydrogelPMMA Chamber

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