CE material

Training CE material

On this page you find teaching materials for the module "Contrast Enhancement in Light Microscopy"

General Resources


Web resources

The tutorial is presented in one piece, so that it can be viewed in logical order, but its components may be viewed separately by reference to the time-points quoted.
1. Conjugate Planes in the microscope (0 – 21:00 min)
The optical system of a microscope embodies two series of planes which are conjugate – optically linked, as images one of another. These are the ‘Field’ set of planes consisting of the object and its images, and the ‘Aperture’ set of planes which includes the light source (lamp filament) and it subsequent images. These planes are demonstrated here using a specially assembled microscope and video camera system.
2. Diffraction and the microscope image (21:00 – 50:51 min)
The most important function of a microscope is resolution – the ability to handle information about fine detail – and this is limited by the phenomenon of diffraction. Two factors are involved – the wavelength of the light, and the Numerical Aperture of the objective lens. The importance of these two factors was demonstrated in 1873 by Ernst Abbe in his ‘Theory of the Microscope’, and illustrated by a series of experiments. These experiments are demonstrated here, again using the specially-adapted microscope. Understanding this section will benefit from having followed section 1.
3. Dark-field Microscopy (51:41 – 59:40 min)
The second important function of a microscope is to provide contrast in the images of transparent and otherwise invisible objects. Dark-field (or Dark-ground) microscopy offers a simple and inexpensive method of enhancing contrast. It is explained here as a sequel to the Diffraction Experiments, using the same demonstration equipment, and should therefore be viewed following sections 1 & 2.
4. Phase-contrast Microscopy (59:40 min)
This section demonstrates the principle of a second system of enhancing visibility of transparent objects, Phase Contrast. Again this is demonstrated using the equipment as in earlier sections, and it should therefore be viewed following these.
The video was enabled by German Research Foundation (DFG) core facility funding of the Biopolis Dresden Imaging Platform (BioDIP) and has been recorded by Christian Spataro.
Screenshot BioDIP contrasts enhancement, Britta Schroth-Diez
Image: Screenshot BioDIP contrast enhancement (Author: Britta Schroth-Diez)

Polarization and Birefrigence

  • Polarization can be nicely demonstrated using two polarization filters:
The video Light extinction by crossed polars on BioDIP you tube channel shows the effect of crossing two polarization filters on light passing through them. When the two polarizers are oriented perpendicular to each other, light gets extinct.
  • Birefringence can be nicely demonstrated using a calcite crystal and a paper with one black spot:
The video Birefringence on BioDIP you tube channel shows the effect of looking at a black spot through a rotating calcite crystal. There are two spots visible of which one is rotating around the other one. (A nice addition to this demonstration is to rotate a polarization filter on top of the calcite crystal. This will make the spots disappear alternately depending on the polarization direction.)