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Related Concept Videos

Total Internal Reflection Fluorescence Microscopy01:05

Total Internal Reflection Fluorescence Microscopy

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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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Attenuated Total Reflectance (ATR) Infrared Spectroscopy: Overview01:13

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Attenuated total reflectance (ATR) infrared spectroscopy is a powerful analytical technique used to study the composition of materials. It is widely employed in chemistry, materials science, forensic science, and other fields where sample characterization is required. ATR has several advantages over traditional transmission IR spectroscopy, including the requirement of little to no sample preparation and the ability to analyze a wide range of samples.
The ATR process begins by directing a beam...
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Super-resolution Fluorescence Microscopy01:37

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Super-resolution fluorescence microscopy (SRFM) provides a better resolution than conventional fluorescence microscopy by reducing the point spread function (PSF). PSF is the light intensity distribution from a point that causes it to appear blurred. Due to PSF, each fluorescing point appears bigger than its actual size, and it is the PSF interference of nearby fluorophores that causes the blurred image. Various approaches to achieving higher resolution through SRFM have recently been...
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Related Experiment Video

Updated: Sep 1, 2025

Simultaneous Interference Reflection and Total Internal Reflection Fluorescence Microscopy for Imaging Dynamic Microtubules and Associated Proteins
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Total Internal Reflection Fluorescence (TIRF) Microscopy.

Kenneth N Fish1

  • 1Department of Psychiatry, University of Pittsburgh School of Medicine, Pittsburgh, Pennsylvania.

Current Protocols
|August 16, 2022
PubMed
Summary
This summary is machine-generated.

Total internal reflection fluorescence microscopy (TIRFM) uses an evanescent wave to excite fluorophores near surfaces. This technique is ideal for studying neuronal plasma membrane receptors with high spatial resolution.

Keywords:
axial resolutionfluorescence microscopylive cell imagingneuronsreceptor trafficking

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Area of Science:

  • Biophysics
  • Optical Microscopy
  • Cell Biology

Background:

  • Total internal reflection fluorescence microscopy (TIRFM) is an optical technique for selective excitation of fluorophores.
  • It utilizes an evanescent wave generated at a solid-liquid interface.
  • This wave confines excitation to a thin layer near the interface, enabling optical sectioning.

Purpose of the Study:

  • To provide an overview of Total internal reflection fluorescence microscopy (TIRFM).
  • To discuss the history, optical theory, and hardware configurations of TIRFM.
  • To detail experimental methods for studying plasma membrane receptors in neurons using TIRFM.

Main Methods:

  • The principle of total internal reflection at a solid-liquid interface is employed.
  • An evanescent electromagnetic wave is generated, decaying exponentially with distance.
  • This wave selectively excites fluorophores within a few hundred nanometers of the interface.

Main Results:

  • TIRFM allows for high-resolution imaging of events near cell surfaces.
  • The technique provides optical sectioning capabilities, reducing background fluorescence.
  • It enables detailed study of receptor dynamics at the plasma membrane.

Conclusions:

  • TIRFM is a powerful tool for visualizing molecular events at the cell membrane.
  • Understanding TIRFM principles and methods is crucial for its effective application.
  • This technique offers significant advantages for neuroscience research, particularly in studying neuronal receptors.