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With an optical microscope having a high numerical aperture and using oil immersion, the best possible resolution is 200 nm corresponding to a magnification of around 1200×. Without oil immersion, the maximum usable magnification is around 800×.
There is a diffraction-limited resolution depending on incident wavelength; in visible range, the resolution of optical microscopy is limited to approximately 0.2 micrometres (see: microscope) and the practical magnification limit to ~1500x. [13] Out-of-focus light from points outside the focal plane reduces image clarity. [14]
Some microscopes make use of oil-immersion objectives or water-immersion objectives for greater resolution at high magnification. These are used with index-matching material such as immersion oil or water and a matched cover slip between the objective lens and the sample. The refractive index of the index-matching material is higher than air ...
Spectral precision distance microscopy (SPDM) is a family of localizing techniques in fluorescence microscopy which gets around the problem of there being many sources by measuring just a few sources at a time, so that each source is "optically isolated" from the others (i.e., separated by more than the microscope's resolution, typically ~200 ...
[1] [2] A fluorescence microscope is any microscope that uses fluorescence to generate an image, whether it is a simple set up like an epifluorescence microscope or a more complicated design such as a confocal microscope, which uses optical sectioning to get better resolution of the fluorescence image. [3]
Fluorescence and confocal microscopes operating principle. Confocal microscopy, most frequently confocal laser scanning microscopy (CLSM) or laser scanning confocal microscopy (LSCM), is an optical imaging technique for increasing optical resolution and contrast of a micrograph by means of using a spatial pinhole to block out-of-focus light in image formation. [1]
The practical limit to magnification with a light microscope is around 1300×. Higher magnifications are possible, but it becomes increasingly difficult to maintain image clarity as the magnification increases. [17] Bright-field microscopes have low apparent optical resolution due to the blur of out-of-focus material;
In a dry objective or condenser, this gives a maximum NA of 0.95. In a high-resolution oil immersion lens, the maximum NA is typically 1.45, when using immersion oil with a refractive index of 1.52. Due to these limitations, the resolution limit of a light microscope using visible light is about 200 nm.
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