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SNARE-mediated Fusion of Single Proteoliposomes with Tethered Supported Bilayers in a Microfluidic Flow Cell Monitored by Polarized TIRF Microscopy
JoVE Journal
Neuroscience
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JoVE Journal Neuroscience
SNARE-mediated Fusion of Single Proteoliposomes with Tethered Supported Bilayers in a Microfluidic Flow Cell Monitored by Polarized TIRF Microscopy
DOI:

10:58 min

August 24, 2016

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Chapters

  • 00:05Title
  • 01:01Preparation of a PDMS Block to Form the Microfluidic Channel
  • 03:17Formation of the Tethered Supported Bilayer
  • 06:18Checking Bilayer Fluidity
  • 07:06Introducing v-SUVs into the Microfluidic Channels
  • 07:59Observing the Fusion of Single v-SUVs and the SBL
  • 08:42Results: Docking and Fusion Events Described by Lipid and Content Release
  • 10:26Conclusion

Summary

Automatic Translation

Here, we present a protocol to detect single, SNARE-mediated fusion events between liposomes and supported bilayers in microfluidic channels using polarized TIRFM, with single molecule sensitivity and ~15 msec time resolution. Lipid and soluble cargo release can be detected simultaneously. Liposome size, lipid diffusivity, and fusion pore properties are measured.

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