The optical microscope is an ancient and young scientific tool. It has a history of 300 years since its birth. The optical microscope has a wide range of uses. For example, in biology, chemistry, physics, astronomy, etc., it is inseparable from the microscope in some scientific research work.

According to different application purposes, microscopes can be roughly classified into four categories: biological microscopes, metallographic microscopes, stereo microscopes, and polarizing microscopes. As the name implies, biological microscopes are mainly used in biomedicine, and the observation objects are mostly transparent or translucent micro-bodies; metallographic microscopes are mainly used to observe the surface of opaque objects, such as the metallographic structure and surface defects of materials; while stereoscopic microscopes magnify micro-objects, they also make objects and images in the same direction relative to the human eye, and have a sense of depth, which is in line with people's conventional visual habits; polarizing microscopes use the transmission or reflection characteristics of different materials for polarized light to distinguish different micro-object components. In addition, some special types can also be subdivided. For example, an inverted biological microscope or a culture microscope is a biological microscope mainly used to observe culture through the bottom of a culture vessel; a fluorescence microscope uses the characteristics of certain substances to absorb light with a specific shorter wavelength and emit light with a specific longer wavelength to discover the existence of these substances and determine their content; a comparison microscope can form juxtaposed or superimposed images of two objects in the same field of view to compare the similarities and differences between the two objects.

Traditional optical microscopes are mainly composed of optical systems and their supporting mechanical structures. The optical systems include objective lenses, eyepieces and condenser lenses, all of which are complicated magnifying glasses made of various optical glasses. The objective lens magnifies the specimen, and its magnification Mobject is determined by the following formula: Mobject =Δ∕f'object, where f'object is the focal length of the objective lens, and Δmay be understood as the distance between the objective lens and the eyepiece. The eyepiece magnifies the image formed by the objective lens again, forming a virtual image at 250mm in front of the human eye for observation. This is the most comfortable observation position for most people. The magnification of the eyepiece M=250/f' eye, where f' is the focal length of the eyepiece. The total magnification of the microscope is the product of the objective lens and the eyepiece, that is, M=M object*M eye=Δ*250/f' eye *f; object. It can be seen that reducing the focal length of the objective lens and the eyepiece will increase the total magnification, which is the key to seeing bacteria and other microorganisms with a microscope, and it is also the difference between it and ordinary magnifying glasses.

So, is it conceivable to reduce the f' object f' mesh without limit, so as to increase the magnification, so that we can see more subtle objects? The answer is no! This is because the light used for imaging is essentially a kind of electromagnetic wave, so diffraction and interference phenomena will inevitably occur during the propagation process, just like the ripples on the water surface that can be seen in daily life can go around when encountering obstacles, and two columns of water waves can strengthen or weaken each other when they meet. When the light wave emitted from a point-shaped luminous object enters the objective lens, the frame of the objective lens hinders the propagation of light, resulting in diffraction and interference. After passing through the objective lens, it can no longer gather at one point, but forms a spot of light with a certain size, and there is a series of light rings with weak and gradually weakening intensity on the periphery. We call the central bright spot an Airy disk. When two light-emitting points are close to a certain distance, the two light spots will overlap until they cannot be recognized as two light spots. Rayleigh proposed a judgment standard, thinking that when the distance between the centers of the two light spots is equal to the radius of the Airy disk, the two light spots can be distinguished. After calculation, the distance between the two light-emitting points at this time is e=0.61 In/n.sinA=0.61 In/N.A. In the formula, In is the light wave wavelength, and the light wave wavelength that can be received by the human eye is about 0.4-0.7um, and n is the refractive index of the medium where the light-emitting point is located. For example, in air, n≈1 , in water, n≈1.33, and A is half of the opening angle of the luminescent point to the frame of the objective lens, and N.A is called the numerical aperture of the objective lens. It can be seen from the above formula that the distance between two points that the objective lens can distinguish is limited by the wavelength of light and the numerical aperture. Since the wavelength of the most sensitive human eye is about 0.5um, and the angle A cannot exceed 90 degrees, sinA is always less than 1. The maximum refractive index of the available light-transmitting medium is about 1.5, so the value of e is always greater than 0.2um, which is the minimum limit distance that can be resolved by an optical microscope. Magnify the image through a microscope. If you want to enlarge the object point distance e that can be resolved by the objective lens with a certain N.A value enough to be resolved by the human eye, you need M.e Greater than or equal to 0.15mm, where 0.15mm is the minimum distance between two micro-objects that can be distinguished by the human eye at 250mm in front of your eyes, so M Greater than or equal to (0.15∕0.61)N.A≈500N.A. It is enough to double the magnification, that is, 500N.A Less than or equal to M Less than or equal to 1000N.A, which is a reasonable selection range of the total magnification of the microscope. No matter how large the total magnification is, it is meaningless, because the numerical aperture of the objective lens has limited the minimum resolvable distance, and it is impossible to distinguish smaller object details by increasing the magnification.

Imaging contrast is another key issue in optical microscopes. The so-called contrast refers to the black-and-white contrast or color difference between adjacent parts on the image surface. It is difficult for the human eye to judge the brightness difference below 0.02, but it is slightly more sensitive to the color difference. Some microscope objects, such as biological specimens, have very little brightness difference between the details, and the design and manufacturing errors of the microscope optical system further reduce the imaging contrast and make it difficult to distinguish. At this time, the details of the object cannot be seen clearly.

Over the years, people have worked hard to improve the resolution and imaging contrast of the microscope. With the continuous advancement of computer technology and tools, the theory and methods of optical design have also been continuously improved. Coupled with the improvement of raw material performance, the continuous improvement of technology and detection methods, and the innovation of observation methods, the imaging quality of the optical microscope has approached the perfection of the diffraction limit. People will use specimen staining, dark field, phase contrast, fluorescence, interference, and polarized light. Imaging instruments have come out one after another, and have superior performance in some aspects, but they still cannot compete with optical microscopes in terms of cheapness, convenience, intuition, and especially suitable for research on living organisms. Optical microscopes still firmly occupy their own positions. On the other hand, combined with laser, computer, new material technology, and information technology, the ancient optical microscope is rejuvenating and showing vigorous vitality. Digital microscopes, laser confocal scanning microscopes, near-field scanning microscopes, two-photon microscopes and instruments with various new functions or that can adapt to various new environmental conditions emerge in an endless stream, which further expands the application field of optical microscopes, as examples. How exciting are the microscopic pictures of rock formations uploaded from the Mars rovers! We can fully believe that the optical microscope will benefit mankind with an updated attitude.