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Using Microfluidics and Fluorescence Microscopy to Study the Assembly Dynamics of Single Actin Filaments and Bundles
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Biochimica
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JoVE Journal Biochimica
Using Microfluidics and Fluorescence Microscopy to Study the Assembly Dynamics of Single Actin Filaments and Bundles

Using Microfluidics and Fluorescence Microscopy to Study the Assembly Dynamics of Single Actin Filaments and Bundles

DOI:
10.3791/63891-v

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08:02 min

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May 05, 2022

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, , , ,
1Université Paris Cité, CNRS,Institut Jacques Monod

Capitoli

  • 00:04Introduction
  • 00:40Polydimethylsiloxane (PDMS) Chamber Assembly
  • 01:45Connect Microfluidic Device
  • 03:11Configuring the Setup with Standard Flow Rates and Changing Solution
  • 04:21Basic Single Filament Experiment: Adenosine Diphosphate (ADP)-Actin Barbed End Depolymerization
  • 06:45Results: Polymerization of Actin Filaments from Spectrin-Actin Seeds when Exposed to Different Actin Binding Proteins
  • 07:24Conclusion

Summary

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We present protocols for simple actin filament microfluidic assays, in combination with fluorescence microscopy, that allow one to accurately monitor individual actin filaments in real-time while sequentially exposing them to different protein solutions.

Tags

MicrofluidicsFluorescence MicroscopyActin FilamentsActin BundlesAssembly DynamicsActin-binding ProteinsPDMS ChamberF-bufferPressure ControlAir Bubbles

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