çš„ Principle of phase contrast microscope
â—Ž Light waves have different amplitudes (brightness), wavelength (color), and phase (which can be achieved by fluctuations in light at a certain time).
â—Ž When light passes through an object, such as changes in wavelength and amplitude, it can be observed by people's eyes. This is the reason why a stained specimen can be observed under a normal microscope. In living cells and unstained biological specimens, the refractive index and thickness of the micro-structures of the cells are slightly different. When the light waves pass, the wavelength and amplitude do not change, and only the phase changes (the corresponding difference is the phase difference). And this small change can not be identified by the human eye, so it is difficult to observe under ordinary microscope.
â—Ž Phase contrast microscope can change the phase of direct or diffracted light, and use the diffraction and interference of light to change the phase difference into amplitude difference (light and dark difference), and it also absorbs part of direct light to increase its contrast. It can therefore be used to observe live cells or unstained specimens.
⊙ structural features
â—Ž The main difference between the phase contrast microscope and the ordinary microscope is that the annular diaphragm is used instead of the iris diaphragm, and the objective lens with the phase plate (usually marked with PH) is used instead of the ordinary objective lens with a shaft adjustment. Telescope.
â—Ž The annular diaphragm is a diaphragm formed by annular holes of different sizes, and their diameter and hole width are matched with different objective lenses.
â—Ž The phase plate is mounted on the back focal plane of the objective lens. The phase plate is equipped with an absorption film that absorbs light and a phase film that delays the phase. In addition to delaying the phase of direct or diffracted light, it also absorbs light to change the brightness.
â—Ž The telescope is used to adjust the uranium. When the phase contrast microscope is used, the center of the annular aperture below the condenser and the optical axis of the objective lens should be completely in line. It is necessary to adjust the bright ring of the pupil and the annular ring of the phase plate to coincide (conjugate overlap) to play the phase contrast microscope. Performance. Otherwise, the optical path of the direct or diffracted light is disordered, and the light to be absorbed cannot be absorbed. The light wave of the delayed phase cannot be delayed, and the effect of the phase contrast microscope is lost.
⊙ phase difference accessory
⊙How to use
(1) Replacement installation of phase difference device
Remove the concentrator used in the ordinary microscope, mount the ring diaphragm on the concentrator bracket, and place the green filter on it. It can absorb red and blue light and illuminate the monochromatic light with a small wavelength range. And has an endothermic effect, which can achieve good results in phase difference observation. Then unscrew the ordinary objective lens from the converter and replace it with the phase difference objective lens.
(2) Focusing
Turn on the light source, rotate the concentrator turntable, and align the “0†with the marking hole to make the ordinary concentrator part enter the optical path. First use the low-magnitude phase difference objective lens to perform light focusing and focusing according to the ordinary microscope operation method. Rotate the annular diaphragm so that the diameter and hole width of the diaphragm are compatible with the phase difference objective used. For example, if the phase difference objective is 40X, the aperture of the 40X mark hole is applied.
(3) Uranium adjustment
Pull out the eyepiece, insert the uranium telescope, and observe it from the telescope inward, and fix the outer cylinder with your left hand. Rotate the telescope inner cylinder with your right hand to raise it. When you focus on the focus, you can see the bright ring of the ring diaphragm. The dark ring of the phase plate, at which point the telescope can be fixed. Then raise and lower the concentrator and adjust the spiral underneath to make the size of the bright ring coincide with the dark ring, then adjust the adjustment knob on the ring diaphragm concentrator left and right to make the two rings completely coincide.
(4) Observation
Change back to the eyepiece and observe as usual. When replacing phase difference objectives of different magnifications, use a matching annular diaphragm each time.
⊙ phase difference adjustment
⊙Use
Used to observe the morphology of living cells in tissue culture. Living cells are colorless and transparent, and it is difficult to distinguish cell contours and structures under light microscopy. In tissue culture studies, inverted phase contrast microscopy is commonly used.
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