In the development of the microscope, the invention of phase contrast mirror observation is an important achievement in modern microscope technology. We know that the human eye can only distinguish between the wavelength (color) and amplitude (brightness) of the light wave. For colorless and transparent biological specimens, when the light passes, the wavelength and amplitude do not change much, and it is difficult to observe the specimen in the bright field observation.
The phase contrast microscope uses the optical path difference of the observed object to observe, that is, the interference phenomenon of the light is used to change the phase difference that is indistinguishable to the human eye into a resolvable amplitude difference, even if the colorless and transparent substance can be clearly seen. This greatly facilitates the observation of living cells, so phase contrast observation is widely used in inverted microscopes.
The basic principle of the phase contrast microscope is to change the optical path difference of the visible light transmitted through the specimen into an amplitude difference, thereby improving the contrast between various structures and making the various structures clearly visible. After the light passes through the specimen, it refracts, deviates from the original optical path, and is delayed by 1/4λ (wavelength). If it increases or decreases by 1/4λ, the optical path difference becomes 1/2λ, and the two beams of light combine with the rear interference. Strengthen, increase or decrease the amplitude, and increase the contrast. Structurally, phase contrast microscopy differs from ordinary optical microscopy in that it has two special features:
- The annular diaphragm is located between the light source and the concentrator, and functions to form a hollow light cone through the light of the concentrator to collect on the specimen.
- The phase plate has a phase plate coated with magnesium fluoride in the objective lens, which can delay the phase of direct or diffracted light by 1/4λ, and is divided into two types:
?? A + phase plate: the direct light is delayed by 1/4 λ, the two sets of light waves and the back-axis light waves are added, the amplitude is increased, the specimen structure is brighter than the surrounding medium, forming a bright contrast (or negative contrast).
‚B+ phase plate: The diffracted light is delayed by 1/4λ, the two sets of light waves and the back-axis light wave are subtracted, the amplitude becomes smaller, and a dark contrast (or positive contrast) is formed, and the structure is darker than the surrounding medium.
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