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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 both cases the numerical aperture of the objective is 1.49 and the refractive index of the medium 1.52. The wavelength of the emitted light is assumed to be 600 nm and, in case of the confocal microscope, that of the excitation light 500 nm with circular polarization. A section is cut to visualize the internal intensity distribution.
Full-spectrum photography is a subset of multispectral imaging, defined among photography enthusiasts as imaging with consumer cameras the full, broad spectrum of a film or camera sensor bandwidth. In practice, specialized broadband/full-spectrum film captures visible and near infrared light, commonly referred to as the " VNIR ".
Cells loaded with an indicator, or expressing it in the case of a GECI, [42] can be viewed using a fluorescence microscope and captured by a Scientific CMOS (sCMOS) [43] camera or CCD camera. Confocal and two-photon microscopes provide optical sectioning ability so that calcium signals can be resolved in microdomains such as dendritic spines or ...
By virtue of the linearity property of optical non-coherent imaging systems, i.e., . Image(Object 1 + Object 2) = Image(Object 1) + Image(Object 2). the image of an object in a microscope or telescope as a non-coherent imaging system can be computed by expressing the object-plane field as a weighted sum of 2D impulse functions, and then expressing the image plane field as a weighted sum of the ...
Köhler illumination is a method of specimen illumination used for transmitted and reflected light (trans- and epi-illuminated) optical microscopy.Köhler illumination acts to generate an even illumination of the sample and ensures that an image of the illumination source (for example a halogen lamp filament) is not visible in the resulting image.
Fluorescence lifetimes can be determined in the frequency domain by a phase-modulation method. The method uses a light source that is pulsed or modulated at high frequency (up to 500 MHz) such as an LED, diode laser or a continuous wave source combined with an electro-optic modulator or an acousto-optic modulator. The fluorescence is (a ...
By 1978, the first theoretical ideas had been developed to break the Abbe limit, which called for using a 4Pi microscope as a confocal laser-scanning fluorescence microscope where the light is focused from all sides to a common focus that is used to scan the object by 'point-by-point' excitation combined with 'point-by-point' detection. [14]
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