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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]
In confocal laser-scanned microscopes, the full-width half-maximum (FWHM) of the point spread function is often used to avoid the difficulty of measuring the Airy disc. [1] This, combined with the rastered illumination pattern, results in better resolution, but it is still proportional to the Rayleigh-based formula given above.
A 1951 USAF resolution test chart is a microscopic optical resolution test device originally defined by the U.S. Air Force MIL-STD-150A standard of 1951. The design provides numerous small target shapes exhibiting a stepped assortment of precise spatial frequency specimens.
Confocal microscopy: Fluorescence, scattering 0.2 3-20 0.3-3 10-100 Two-photon microscopy: Fluorescence 0.5-1.0 1-10 0.3-3 10-100 Optical coherence tomography: 300 T Optical scattering 1-2 0.5-10 1-10 20-4.000 Scanning laser acoustic microscopy: 300 M Ultrasonic scattering 1-2 20 20 10 Acoustic microscopy: 50 M Ultrasonic scattering 20 20-100 ...
A 4Pi microscope is a laser scanning fluorescence microscope with an improved axial resolution.With it the typical range of the axial resolution of 500–700 nm can be improved to 100–150 nm, which corresponds to an almost spherical focal spot with 5–7 times less volume than that of standard confocal microscopy.
Laser scanning is the controlled deflection of laser beams, visible or invisible. [1] Scanned laser beams are used in some 3-D printers, in rapid prototyping, in machines for material processing, in laser engraving machines, in ophthalmological laser systems for the treatment of presbyopia, in confocal microscopy, in laser printers, in laser shows, in Laser TV, and in barcode scanners.
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