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Related Concept Videos

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.
Protein Dynamics in Living Cells01:19

Protein Dynamics in Living Cells

Different fluorescence-based techniques are used to study the protein dynamics in living cells. These techniques include FRAP, FRET, and PET.
Fluorescent recovery after photobleaching (FRAP) is a fluorescent-protein-based detection technique used to quantify protein movement rates within the cell. This method exposes a small portion of the cell to an intense laser beam. The laser beam causes permanent photobleaching of the fluorophore-tagged proteins in the exposed region. As the bleached...
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.
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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Related Experiment Video

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Simultaneous Multicolor Imaging of Biological Structures with Fluorescence Photoactivation Localization Microscopy
12:51

Simultaneous Multicolor Imaging of Biological Structures with Fluorescence Photoactivation Localization Microscopy

Published on: December 9, 2013

Photoactivatable fluorescent proteins for diffraction-limited and super-resolution imaging.

Jennifer Lippincott-Schwartz1, George H Patterson

  • 1Cell Biology and Metabolism Program, NICHD, NIH Bethesda, MD, USA. jlippin@helix.nih.gov

Trends in Cell Biology
|October 20, 2009
PubMed
Summary

Photoactivatable fluorescent proteins (PA-FPs) enable high-contrast imaging by switching fluorescence states. This review covers PA-FP types and their use in advanced biological imaging techniques.

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Last Updated: Jun 19, 2026

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Published on: December 9, 2013

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Super-Resolution Imaging and Shared Management: A Protocol for Confocal Microscopy with Multiplex Detection

Published on: February 24, 2026

Area of Science:

  • Biochemistry
  • Molecular Biology
  • Microscopy

Background:

  • Photoactivatable fluorescent proteins (PA-FPs) are crucial tools for generating high contrast in biological imaging.
  • Recent advancements have led to diverse PA-FP types with varied optical properties.

Purpose of the Study:

  • To review the expanding array of PA-FPs.
  • To discuss their advantages and disadvantages.
  • To highlight their applications in novel imaging methodologies.

Main Methods:

  • Survey of existing literature on PA-FPs.
  • Analysis of PA-FP properties (fluorescence, activation).
  • Review of imaging techniques utilizing PA-FPs.

Main Results:

  • Development of PA-FPs with green/red fluorescence or green-to-red conversion.
  • Emergence of reversibly switchable PA-FPs.
  • Application in advanced imaging like pulse-chase labeling and super-resolution microscopy.

Conclusions:

  • PA-FPs offer significant advantages for biological research.
  • Novel imaging techniques with PA-FPs provide critical insights into cellular processes.
  • Continued development of PA-FPs promises further advancements in biological imaging.