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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.
Super-resolution Fluorescence Microscopy01:37

Super-resolution Fluorescence Microscopy

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 developed.
Confocal Fluorescence Microscopy01:16

Confocal Fluorescence Microscopy

Confocal microscopy is an advanced microscopic technique. The prime advantage of the confocal microscope over other microscopy techniques is its ability to block the out-of-focus light from the illuminated samples using pinholes. It is widely used with fluorescence optics to obtain high-resolution, sharp contrast images. Unlike optical microscopes, confocal microscopes use a focused beam of light laser to scan the entire sample surface at different z-planes. These microscopes are, therefore,...

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Updated: Jun 24, 2026

High-Throughput Total Internal Reflection Fluorescence and Direct Stochastic Optical Reconstruction Microscopy Using a Photonic Chip
14:09

High-Throughput Total Internal Reflection Fluorescence and Direct Stochastic Optical Reconstruction Microscopy Using a Photonic Chip

Published on: November 16, 2019

Fast two-dimensional standing-wave total-internal-reflection fluorescence microscopy using acousto-optic deflectors.

Olga Gliko1, William E Brownell, Peter Saggau

  • 1Department of Otorhinolaryngology-Head and Neck Surgery, Baylor College of Medicine, Houston, TX 77030, USA.

Optics Letters
|March 14, 2009
PubMed
Summary
This summary is machine-generated.

This study introduces a novel 2D standing-wave total-internal-reflection fluorescence microscopy technique. It achieves sub-100 nm resolution and axial selectivity for real-time imaging of subresolution structures in live biological samples.

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Area of Science:

  • Biophysics
  • Optical Microscopy
  • Cell Biology

Background:

  • Total-internal-reflection fluorescence microscopy (TIRFm) is crucial for live-cell imaging.
  • Achieving high axial resolution (<100 nm) remains a challenge in TIRFm.

Purpose of the Study:

  • To develop a 2D standing-wave (SW) based TIRFm technique.
  • To enhance lateral resolution and axial selectivity for subresolution structure imaging.

Main Methods:

  • Generating 2D SW patterns using two interfering beams coupled via an objective lens.
  • Controlling SW period, orientation, and phase using acousto-optic deflectors.
  • Utilizing an evanescent SW pattern for axial confinement.

Main Results:

  • Achieved lateral resolution improvement of 100 nm.
  • Demonstrated axial selectivity of less than 100 nm.
  • Enabled real-time imaging of subresolution structures.

Conclusions:

  • The developed 2D SW TIRFm technique offers significant resolution and selectivity improvements.
  • This method is suitable for studying dynamic biological processes at the glass-water interface.
  • Provides a new tool for high-resolution live-cell imaging near surfaces.