Several issues in the use of phase contrast microscopy:
(1) Phase reversal when n '
(2) The halo and gradual dimming effect in the imaging process of phase contrast microscopy, when a structure becomes darker due to phase delay, is not a loss of light, but a result of light redistribution on the image plane. Therefore, light that clearly disappears in dark areas will appear as a bright halo around darker objects. This is a disadvantage of phase contrast microscopy, which hinders the observation of fine structures. When the annular aperture is very narrow, the halo phenomenon is more severe. Another phenomenon of phase contrast microscopy is the dimming effect, which refers to a decrease in contrast at the edges of a larger area with the same phase delay observed during phase contrast observation.
(3) The influence of sample thickness When observing the difference, the thickness of the sample should be 5 μ m or thinner. When using thicker samples, the upper layer of the sample is clear, while the deeper layer will be blurry and produce phase shift interference and light scattering interference.
(4) The influence of cover glass and slide on the sample must be covered with a cover glass, otherwise the bright ring of the annular aperture and the dark ring of the phase plate are difficult to overlap. Differential observation also has high requirements for the quality of the glass of the slide and cover glass. When there are scratches, uneven thickness, or uneven unevenness, it can cause distortion of the bright ring and phase interference. In addition, if the glass slide is too thick or too thin, it will cause the annular aperture to become larger or smaller.
At present, optical microscopes have evolved from traditional biological microscopes to various types of specialized microscopes. According to their imaging principles, they can be divided into:
① Geometric optical microscope: including biological microscope, falling light microscope, inverted microscope, metallographic microscope, dark field microscope, etc.
② Physical optical microscope: including phase contrast microscope, polarization microscope, interference microscope, phase contrast polarization microscope, phase contrast interference microscope, phase contrast fluorescence microscope, etc.
③ Information conversion microscopes: including fluorescence microscopes, microspectrometers, image analysis microscopes, acoustic microscopes, photographic microscopes, television microscopes, etc.
List several uses of microscopes: a Biological microscope: Generally speaking, microscopes can be divided into stereo microscopes and biological microscopes. Due to different purposes and requirements, many branches have emerged, but the basic principles remain the same. Polarization, phase contrast, transmission, and falling light are still classified as biological microscopes. Stereoscopic microscope, also known as anatomical microscope, solid microscope, and stereo microscope, is a versatile microscope. It is easy to operate, has low requirements for specimens, has a long working distance, and has a strong sense of three dimensionality when observing. It can be used to observe physical objects or perform some operations on specimens while observing. Instead of slicing the specimen like a biological microscope, slicing requires corresponding technology and equipment. Therefore, stereo microscopes have a wide range of applications in fields such as microelectronics, precision instrument assembly and maintenance, and micro engraving. Widely used in anatomy and microsurgery in the fields of biology and medicine (currently classified as surgical microscopes), the light source used in biology and medicine can only be a cold light source (fiber optic); Used in industry for observation, assembly, inspection, and other work of small parts and integrated circuits. Metallographic microscope: Many people like to write it as "metallographic microscope". Metallographic microscope is a microscope specifically used to observe the metallographic structure of opaque objects such as metals and minerals. These opaque objects cannot be observed in a regular transmission microscope, so the main difference between them and a regular microscope is that the former uses reflected light, while the latter uses transmitted light for illumination. In a metallographic microscope, the illumination beam is directed from the objective lens to the surface of the observed object, reflected by the surface, and then returned to the objective lens for imaging. This reflective lighting method is also widely used in the detection of integrated circuit silicon wafers.
