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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.
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,...
Two-Dimensional Microscopy in Microbiology01:29

Two-Dimensional Microscopy in Microbiology

Two-dimensional (2D) microscopy encompasses a range of optical techniques that capture images within a single focal plane, offering detailed representations of microscopic structures. These techniques are essential in biological and medical research, enabling the visualization of cellular and subcellular structures with different levels of contrast and specificity.There are several major types of 2D microscopy, each with strengths and applications.Bright-Field MicroscopyBright-field microscopy...
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.
Imaging Biological Samples with Optical Microscopy01:18

Imaging Biological Samples with Optical Microscopy

Optical microscopy uses optic principles to provide detailed images of samples. Antonie van Leeuwenhoek designed the first compound optical microscope in the 17th century to visualize blood cells, bacteria, and yeast cells. In 1830, Joseph Jackson Lister created an essentially modern light microscope. The 20th century saw the development of microscopes with enhanced magnification and resolution.
In optical microscopy, the specimen to be viewed is placed on a glass slide and clipped on the stage...
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...

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Conducting Multiple Imaging Modes with One Fluorescence Microscope
08:32

Conducting Multiple Imaging Modes with One Fluorescence Microscope

Published on: October 28, 2018

Fluorescence microscopy beyond the diffraction limit.

Mike Heilemann1

  • 1Applied Laser Physics and Laser Spectroscopy, Physics Department, Bielefeld University, Universitätsstrasse 25, 33615 Bielefeld, Germany. heileman@physik.uni-bielefeld.de

Journal of Biotechnology
|March 30, 2010
PubMed
Summary
This summary is machine-generated.

New far-field microscopy techniques overcome the diffraction barrier, enabling subdiffraction resolution in biological samples. These advanced methods reveal intricate details of biomolecular structures and dynamics in living cells.

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Area of Science:

  • Biophysics
  • Optical Microscopy
  • Cell Biology

Background:

  • Light microscopy is limited by the diffraction barrier, restricting spatial resolution.
  • Recent advancements have introduced fluorescence microscopy techniques that surpass this diffraction limit.
  • Far-field microscopic methods are crucial for studying complex biological systems like cells and tissues.

Purpose of the Study:

  • To introduce key concepts of far-field microscopy with subdiffraction resolution.
  • To discuss the underlying physical principles of these advanced microscopy techniques.
  • To provide practical considerations for applying these methods in biological research.

Main Methods:

  • Review of various far-field fluorescence microscopy techniques.
  • Explanation of the physics enabling subdiffraction resolution.
  • Discussion of practical implementation in biological samples.

Main Results:

  • Emergence of fluorescence microscopy methods bypassing the diffraction barrier.
  • Demonstration of subdiffraction spatial resolution in far-field techniques.
  • Application of these techniques for novel insights into biomolecular organization and dynamics.

Conclusions:

  • Far-field microscopy offers unprecedented resolution for biological imaging.
  • These techniques provide valuable insights into molecular structures and processes in living cells.
  • Understanding the concepts and practicalities is key for effective application.