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Single-Molecule Localization Microscopy and Tracking with a Fluorescent Mechanosensitive Probe.

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  • 1Department of Physical Chemistry, University of Geneva, 1211 Geneva, Switzerland.

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|August 9, 2024
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Summary
This summary is machine-generated.

The Flipper-TR probe enables single-molecule super-resolution imaging of cell membranes, resolving both ordered and disordered lipid regions. This advancement allows for detailed analysis of membrane composition through molecular diffusion tracking.

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Area of Science:

  • Biophysics
  • Cell Biology
  • Optical Imaging

Background:

  • Flipper fluorescent probes report cellular mechanical forces via fluorescence lifetime.
  • Flipper-CF3 probe, designed for super-resolution microscopy, proved too dim for single-molecule detection in disordered membranes.

Purpose of the Study:

  • To evaluate the Flipper-TR probe for single-molecule super-resolution imaging.
  • To assess Flipper-TR's capability in resolving different lipid membrane types.
  • To investigate Flipper-TR's potential for distinguishing membrane compositions via diffusion analysis.

Main Methods:

  • Single-molecule super-resolution fluorescence microscopy.
  • Tracking of individual Flipper-TR probe molecules in lipid bilayers.
  • Analysis of probe diffusion coefficients to infer membrane composition.

Main Results:

  • Flipper-TR is compatible with single-molecule super-resolution imaging.
  • Flipper-TR resolves both liquid ordered and liquid disordered membranes in giant unilamellar vesicles.
  • Probe diffusion coefficients differentiate membranes of varying lipid composition.
  • Brightness differences between Flipper-TR and Flipper-CF3 stem from their optical properties.

Conclusions:

  • Flipper-TR is a viable tool for single-molecule super-resolution imaging of membrane tension.
  • The Flipper-TR scaffold is adaptable for various super-resolution applications, including targeted derivatives.