Dark-field microscopy (dark microscope) describes microscopy methods, both in light and electron microscopes, which exclude the untouched light from the image. As a result, the fields around the specimen (ie, where there are no specimens to spread the file) are generally dark.
Video Dark-field microscopy
Aplikasi mikroskopi ringan
In an optical microscope, the dark plane describes the lighting technique used to increase contrast in an unstained sample. It works by illuminating the sample with light that will not be collected by the objective lens and thus will not form part of the image. It produces a classic dark, almost black, background with a bright object on it.
Light path
The steps are illustrated in the picture where inverted microscope is used.
- Light enters the microscope for sample illumination.
- Custom-sized disks, stop settings (see picture), block some light from the light source, leaving the outer circle of illumination. The broad phase of the aulus can also be replaced by low magnification.
- The condenser lens focuses the light toward the sample.
- Light enters the sample. Most are transmitted instantly, while some are scattered from the sample.
- The scattered light enters the objective lens, while the instantly transmitted just misses the lens and is not collected because of the direct illumination block (see picture ).
- Only the scattered lights illuminate to produce the image, while the transmitted light is directly removed.
Advantages and disadvantages
Dark field microscopy is a very simple yet effective technique suitable for use involving live and non-stained biological samples, such as swabs of tissue or individual cultures, single-celled, single-celled organisms. Considering the simplicity of the settings, the image quality obtained from this technique is very impressive.
The main limitation of dark field microscopy is the low light level seen in the final image. This means that the sample should be very strongly illuminated, which can cause damage to the sample. Dark field microscopy techniques are almost entirely free of artifacts, due to the nature of the process. However, dark-field image interpretation should be done with caution, since the common dark features of bright field microscopy images may not be visible, and vice versa.
While dark field images may first appear negative from bright field images, different effects are seen in each. In bright field microscopy, features can be seen where the shadows are laid out on the surface by incident light or some surface less reflective, possibly by a hole or scratch. Lifted features that are too smooth to produce the shadow will not appear in the bright field image, but the light reflecting on the side of the feature will be visible in dark field images.
- Comparison of transillumination techniques used to produce contrast in tissue paper samples (1.559? m/pixel when viewed at full resolution)
Use in computing
Dark field microscopes have recently been used in computer mouse pointer devices, to allow the optical mouse to work on transparent glass by imaging microscopic defects and dust on its surface.
Dark field microscopy combined with hyperspectral imagery
When combined with hyperspectral imaging, dark field microscopy becomes a powerful tool for the characterization of embedded nanomaterials in cells. In a recent publication, Patskovsky et al. using this technique to study the attachment of gold nanoparticles (AuNPs) that target CD44 cell cancer.
Maps Dark-field microscopy
Transmission electron microscopy applications
Dark field studies in transmission electron microscopy play a strong role in the study of crystals and crystal defects, as well as in individual atomic imaging.
Conventional terrain imagery
Briefly, the imaging involves inclining incident illumination until it is diffracted, rather than incident, the light passes through a small objective hole in the objective lens back in the focal plane. Dark field images, under these conditions, allow one to map the diffracted intensity originating from a collection of diffracted plots as a function of the position projected on the specimen and as a function of the specimen's slope.
In single-crystal specimens, dark field images of a single reflection of a specimen tilted loose from Bragg's condition allow one to "ignite" only lattice defects, such as dislocations or sediments, which bend a set of lattice planes in their environment.. The intensity analysis in the figure can then be used to estimate the amount of bending. In polycrystalline specimens, on the other hand, dark-field images serve to illuminate only the part of the Bragg-reflecting crystal at a particular orientation.
Weak-beam imagery
The weak-beam imagery involves optics similar to the conventional dark field, but the use of the diffracted rays is harmonized rather than the diffracted beam itself. A much higher resolution of the tense area around the defect can be obtained in this way.
Low-angle and high annular dark field imagery
Annular dark-field imaging requires one to form images with electrons diffracted into annular openings centered on, but excluding, untouched rays. For large scattering angles in transmission scanning electron microscopes, this is sometimes called Z -contrast imaging due to increased scattering of high-atom atomic atoms.
Digital dark field analysis
It is a mathematical technique between between the direct and the fourier-transforms to explore images with well-defined periodicities, such as electron microscope lattice images. As with analogue dark-field imaging in a transmission electron microscope, it allows one to "ignite" those objects in the field of view in which the periodicity of interest lies. Unlike analog dark field imagery, it also allows one to map the periodicity of the fourier-phase, and hence the phase gradient, which provides quantitative information on the lattice vector strain.
See also
- Annular dark-field imaging
- Light field microscope
- Wavelet
Footnote
External links
- Nikon - Stereomicroscopy & gt; Darkfield Illumination
- Molecular Expression
- Darkfield Illumination Primer
- Gage SH. 1920. Modern dark-field microscope and history of its development. Transactions of the American Microscopical Society 39 (2): 95-141.
- Dark field and phase contrast microscope (Università ©  © Paris Sud)
Source of the article : Wikipedia
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