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Confocal Microscopy

Protocol
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Part of the Springer Protocols Handbooks book series (SPH)

1. Introduction

Fluorescence microscopy, whether using immunolabeling or expressible markers such as green fluorescent protein (GFP), has become one of the major tools of the cell biologist. In the fluorescence microscope we can identify cellular structures, organelles and macromolecular assemblies with exquisite precision, but because these structures appear bright on a dark background, strongly labeled objects outside the plane of focus can become extremely distracting. In the worst case they can completely swamp fine details that are in focus. The confocal microscope uses an ingenious bit of optical engineering to overcome this problem, and can in fact create fully three-dimensional images of complex specimens.

The principle of confocal optics goes back more than 50 yr, but early implementations were clumsy, and both the mechanical and electronic components then available limited their performance. Most early instruments worked in reflection, not fluorescence, which was useful in...

Keywords

Second Harmonic Generation Point Spread Function Numerical Aperture Lateral Resolution Axial Resolution 
These keywords were added by machine and not by the authors. This process is experimental and the keywords may be updated as the learning algorithm improves.

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Copyright information

© Humana Press, a part of Springer Science+Business Media, LLC 2008

Authors and Affiliations

  • Guy Cox
    • 1
  1. 1.Electron Microscope UnitUniversity of SydneySydneyAustralia

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