Three types of microscope observation
I. Bright field BF (Bright field BF)
Bright field BF is a familiar way of microscopy, which is widely used in pathology and testing for observing stained sections, and all microscopes are capable of performing this function.
Bright field
II. Dark field DF (Dark field DF)
Dark field DF is actually dark field illumination. It is different from bright field in that it does not directly observe the illuminated light, but the light reflected or diffracted from the object being examined. As a result, the field of view becomes a dark background, while the examined object appears as a bright image.
The principle of dark field of view is based on the Tyndall phenomenon in optics, dust in the case of strong light through the direct light, the human eye can not be observed, this is because of the strong light around the cause. If light is directed obliquely at it, the particles seem to increase in size due to the reflection of light and become visible to the human eye.
A special accessory required for dark-field observation is a dark-field spotting scope. It is characterised by not allowing the light beam to pass through the examined object from bottom to top, but by changing the path of the light so that it is directed obliquely towards the examined object, so that the illuminating light does not enter directly into the objective lens, and a bright image is formed by using the reflected or diffracted light from the surface of the examined object. The resolution of dark-field observation is much higher than bright-field observation, * up to 0.02-0.004
Darkfield
III. Phase contrast PH
In the development of optical microscopy, the successful invention of phase contrast PH is an important achievement in modern microscopy technology. As we know, the human eye can only distinguish the wavelength (colour) and amplitude (brightness) of light waves, for colourless and bright biological specimens, when the light passes through, the wavelength and amplitude do not change much, and it is difficult to observe the specimen in the bright field observation.
Phase contrast microscope uses the difference of the light range of the examined object for microscopic examination, that is, it effectively uses the interference phenomenon of light to change the indistinguishable phase difference of the human eye into the distinguishable amplitude difference, and even colourless and transparent substances can become clearly visible. This greatly facilitates the observation of living cells, so phase contrast microscopy is widely used in inverted microscopes.
The basic principle of phase contrast microscopy is that the difference in optical range of visible light transmitted through a specimen is turned into a difference in amplitude, thus increasing the contrast between various structures and making them visible. The light is refracted through the specimen and deviates from the original light path, while being delayed by 1/4λ (wavelength). If the 1/4λ is increased or decreased again, the optical range difference becomes 1/2λ, and the interference between the two beams of photosynthesis is strengthened after the axes of the two beams are interfered with, and the amplitude increases or decreases, thus improving the contrast. In the structure, phase contrast microscope has two special features which are different from ordinary optical microscope:
1. annular diaphragm (annulardiaphragm) is located between the light source and the condenser, the role is to make the light through the condenser to form a hollow cone of light, focusing on the specimen.
2. phase plate (annularphaseplate) in the objective lens coated with magnesium fluoride phase plate, direct or diffracted light can be delayed phase 1/4λ. There are two kinds:
1.A phase plate: the direct light delayed 1/4 λ, two groups of light waves co-axial light wave addition, amplitude increase, the specimen structure than the surrounding medium is brighter, the formation of bright contrast (or negative contrast).
2.B phase plate: the diffracted light is delayed by 1/4 λ, the two groups of light waves after the merging of the axis of the light wave is reduced, the amplitude becomes smaller, the formation of dark contrast (or positive contrast), the structure is darker than the surrounding medium.
