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Mass-Sensitive Particle Tracking to Characterize Membrane-Associated Macromolecule Dynamics
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Biochemistry
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JoVE Journal Biochemistry
Mass-Sensitive Particle Tracking to Characterize Membrane-Associated Macromolecule Dynamics

Mass-Sensitive Particle Tracking to Characterize Membrane-Associated Macromolecule Dynamics

DOI:
10.3791/63583-v

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13:30 min

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February 18, 2022

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, , ,
1Department of Cellular and Molecular Biophysics,Max Planck Institute of Biochemistry, 2Department of Physics,Technical University Munich, 3Department of Cellular Physiology, Biomedical Center (BMC),Ludwig-Maximilians-Universität München

Chapters

  • 00:05Introduction
  • 00:43Cleaning of Microscope Slides
  • 01:48Assembly of Flow Chambers
  • 02:41Supported Lipid Bilayer (SLB) Formation
  • 03:31Imaging
  • 04:31Data Analysis
  • 10:25Results: Native and Ratiometric View of Exemplary Surfaces
  • 11:01Results: Calibration of the Mass-to-Contrast Relation for MSPT Measurements
  • 12:15Results: Deciphering Oligomer States of Membrane-Associated Proteins
  • 12:57Conclusion

Summary

Automatic Translation

Automatic Translation

This protocol describes an iSCAT-based image processing and single-particle tracking approach that enables the simultaneous investigation of the molecular mass and the diffusive behavior of macromolecules interacting with lipid membranes. Step-by-step instructions for sample preparation, mass-to-contrast conversion, movie acquisition, and post-processing are provided alongside directions to prevent potential pitfalls.

Tags

Keywords: Mass-sensitive Particle TrackingMembrane-associated MacromoleculesLabel-freeOligomeric StatesLipid MembranesBiomolecule DynamicsLigand-induced Receptor ComplexesSmall Unilamellar VesiclesFlow ChamberMass Photometer

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