Accu-Scope 3025 Supplemental Instructions

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troubleshooting guide

Accu-Scope 3025 Supplemental Instructions Manual pdf 1 page image

73 Mall Drive • Commack, NY 11725 • www.accu-scope.com • Phone (631) 864-1000 • Fax (631) 543-89003025 MicroscopePhase Contrast – Turret SystemSUPPLEMENTAL INSTRUCTIONSPHASE CONTRAST MICROSCOPYThe normal microscopic object is seen because it has regions of varying density. In normal brightfield illumination acompletely transparent specimen is very difficult to observe in detail because all areas of the specimen are equally dense.Darkfield illumination displays border effects in completely transparent specimens due to edge scattering and diffraction oflight. Polarized light is useful when transparent specimens have directional or crystalline properties.Phase contrast microscopy is a type of illumination system to observe transparent media. This form of illumination isutilized extensively in the study of transparent living cells without the need for staining or fixing while being able to obtaingood image contrast. The light from phase contrast illumination arrives at the user’s eyes at ½ the normal wavelength.This light altering system produces a visible image of an otherwise invisible, transparent specimen.The optical light path necessary for phase contrast is shown in Figure 1. A clear annulus in the focal plane of thecondenser is imaged at infinity by the condenser and then re-imaged by the objective in its rear focal plane. Theundiffracted light passes through this image. It is reduced in intensity and given a one-quarter wave phase shift by meansof an annular phase pattern in the rear focal plane of the objective. These two changes in the undiffracted portion of thebeam simulate the phase and intensity distribution which would be present in the objective focal plane if the specimen haddensity variations rather than refractive index variations. As a result, the image formed by the beam interfering with thediffracted beam simulates that of a specimen having density variations.IMAGE FORMATION BY PHASE CONTRASTAn annular aperture in the diaphragm placed in the focal plane of the substage condenser controls the illumination of thespecimen. The aperture is imaged by the condenser and objective at the rear focal plane or at the exit pupil of theobjective. A phase shifting element, or phase plate, is placed in the image plane. Light passing through the phase alteringpattern acquires a ¼ wave length advance over that diffracted by the object structure and passes through that region ofthe phase plate not covered by the altering pattern. The resultant interference effects of the two portions of light form thefinal image. Altered phase relations in the illumination rays, induced by otherwise invisible elements in the specimen, aretranslated into brightness differences by the phase altering plate.

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