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Related Concept Videos

Total Internal Reflection Fluorescence Microscopy01:05

Total Internal Reflection Fluorescence Microscopy

Total internal reflection fluorescence microscopy or TIRF is an advanced microscopic technique used to visualize fluorophores in samples close to a solid surface with a higher refractive index, such as a glass coverslip. TIRF only allows fluorophores in proximity to the solid surface to be excited. When light from a medium with a lower refractive index (such as air) hits the glass coverslip at a critical angle, the light undergoes total internal reflection stead of passing through the glass.

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Fabrication of Zero Mode Waveguides for High Concentration Single Molecule Microscopy
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Published on: May 12, 2020

Evanescent-wave fluorescence microscopy using symmetric planar waveguides.

Björn Agnarsson1, Saevar Ingthorsson, Thorarinn Gudjonsson

  • 1Department of Physics, Science Institute, University of Iceland, Reykjavik, Iceland.

Optics Express
|April 1, 2009
PubMed
Summary

A novel optical waveguide method enhances fluorescence imaging for biological samples. This technique offers superior image quality and tunability for applications like cell membrane studies.

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Area of Science:

  • Optics and Photonics
  • Biomedical Imaging
  • Biophysics

Background:

  • Evanescent-wave excitation is crucial for high-resolution imaging of biological interfaces.
  • Existing waveguide methods face limitations in image quality and tunability.
  • Advanced optical techniques are needed for detailed studies of cellular interactions.

Purpose of the Study:

  • To introduce a new evanescent-wave fluorescence excitation method using a planar optical waveguide.
  • To demonstrate the advantages of this method for biological sample imaging.
  • To highlight its suitability for various cell-based and surface-binding assays.

Main Methods:

  • A planar optical waveguide was constructed with a core layer supporting light propagation.
  • The waveguide core was symmetrically cladded by the biological sample (aqueous solution) and a matching polymer.
  • Evanescent-wave excitation was generated within the waveguide structure.

Main Results:

  • The described method achieved superior image quality compared to other waveguide-excitation techniques.
  • The evanescent field penetration depth was widely tunable.
  • The system demonstrated compatibility with optical fibers for versatile application.

Conclusions:

  • This novel waveguide-based evanescent-wave excitation method provides a powerful tool for biological imaging.
  • Its advantages in image quality, tunability, and compatibility make it ideal for studying cell membranes and surface interactions.
  • The technique is well-suited for applications in cell culture, including cell-cell and cell-matrix interactions, and monitoring binding events.