Phase Contrast Microscopy

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Here is an article that discusses about the different features of a phase contrast microscope, its function, and comparison from other types of microscopes, and some interactive tutorials and literature references. Phase contrast microscope is described as a contrast-enhancing optical technique that can be utilized to produce high-contrast images of transparent specimens such as living cells, microorganisms, thin tissue slices, lithographic patterns, and sub-cellular particles, like nuclei and other organelles.

The history of phase contrast microscopy was first realized in 1934 by Dutch physicist Frits Zernike. At some stage in his study of diffraction gratings, he valued both that it is necessary to interfere with a reference beam, and that to maximize the contrast achieved with the technique, it is necessary to introduce a phase shift to this reference so that the no-phase-change condition gives rise to completely destructive interference. The phase contrast system utilizes an optical mechanism to translate minute variations in phase into corresponding changes in amplitude, which can be visualized as differences in image contrast. The phase contrast microscope was awarded with the Nobel Prize in Physics, 1953.

The Phase Contrast Microscope functions such as light travels through a medium, contact with this medium causes its amplitude and phase to change, depending on properties of the medium. Changes in amplitude will affect the absorption of light which gives rise to colors when it is wavelength dependent. This difference in phase is not visible to the human eye, so changes in phase are not easily observed, but the changes in phase brings a large amount of information.

Its strong point as a microscope is the ability to examine living cells in their natural state without being killed, fixed, and stained. With this, constant biological processes in live cells can be observed and recorded in high contrast with sharp clarity of minute specimen detail.

The phase contrast microscope is an essential instrument in biological and medical research, especially when dealing with transparent and colorless components in a cell; this makes the transparent object stand out in contrast to its surroundings.

The phase contrast microscope suffers from “halo effect” but with the improvement of the objective phase ring, “apodized phase contrast” was introduced and it allows structures of phase objects having large phase differences to be viewed and photographed with clarity and definition of detail.

The article also relates the comparison between differential interference contrast as the phase contrast produces image intensity values as a function of specimen optical path length magnitude, with very dense regions appearing darker than the background. The phase contrast microscope can come together with a fluorescence microscope for the effects of photo bleaching. And it allows to be added as a component in any brightfield microscope.

You can use the interactive tutorials in the article such as; phase plate/ring alignment, diffracted light in phase contrast microscopy, phase contrast microscope alignment, optical pathways in the phase contrast microscope, phase plate configuration effects on specimen contrast, positive and negative phase contrast, specimen optical path length variations, interaction of light waves with phase specimens, shade-off and halo phase contrast artifacts, apodidized phase contrast, apodized phase plates and specimen contrast, optical sectioning with phase contrast and DIC. And you can view its digital image galleries and/or visit other literature references.

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Here is an article that discusses about the different features of a phase contrast microscope, its function, and comparison from other types of microscopes, and some interactive tutorials and literature references. Phase contrast microscope is described as a contrast-enhancing optical technique that can be utilized to produce high-contrast images of transparent specimens such as living cells, microorganisms, thin tissue slices, lithographic patterns, and sub-cellular particles, like nuclei and other organelles.

The Phase Contrast Microscope functions such as light travels through a medium, contact with this medium causes its amplitude and phase to change, depending on properties of the medium. Changes in amplitude will affect the absorption of light which gives rise to colors when it is wavelength dependent. This difference in phase is not visible to the human eye, so changes in phase are not easily observed, but the changes in phase brings a large amount of information.

Read Original Text here:
http://micro.magnet.fsu.edu/primer/techniques/phasecontrast/phaseindex.html