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Related Experiment Videos

Total internal reflection fluorescence microscopy: application to substrate-supported planar membranes

N L Thompson1, K H Pearce, H V Hsieh

  • 1Department of Chemistry, University of North Carolina, Chapel Hill 27599-3290.

European Biophysics Journal : EBJ
|January 1, 1993
PubMed
Summary
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Total internal reflection fluorescence microscopy (TIRFM) enables studying fluorescent macromolecules binding to model membranes. This review covers methods for analyzing macromolecular motion, organization, and orientation at interfaces.

Area of Science:

  • Biophysics
  • Surface Science
  • Microscopy

Background:

  • Total internal reflection fluorescence microscopy (TIRFM) is a powerful technique for studying molecular interactions at surfaces.
  • Planar model membranes on glass or quartz substrates provide well-defined systems for investigating macromolecular binding.
  • Understanding macromolecular behavior at interfaces is crucial in various biological and material science applications.

Purpose of the Study:

  • To review the applications of TIRFM for studying fluorescent macromolecules that bind to model membranes.
  • To discuss methods for characterizing macromolecular motion, organization, and orientation.
  • To summarize current applications and outline future directions for TIRFM.

Main Methods:

  • Characterization of equilibrium binding using binding curves to determine binding constants.

Related Experiment Videos

  • Analysis of fluorescence photobleaching recovery curves to obtain kinetic rate constants and diffusion coefficients.
  • Measurement of fluorescence intensity versus evanescent field polarization to assess orientational order.

    • Main Results:

    • TIRFM allows for quantitative analysis of macromolecular binding kinetics and thermodynamics.
    • The technique can determine surface diffusion coefficients and orientational order of bound macromolecules.
    • Applications in studying cell-substrate contact regions are highlighted.

    Conclusions:

    • TIRFM is a versatile tool for investigating macromolecular interactions with model membranes.
    • The discussed methods provide comprehensive insights into macromolecular dynamics and organization at interfaces.
    • Future developments in TIRFM hold promise for advancing our understanding of interfacial phenomena.