The relationship between total magnification and resolution of microbiological oil microscopes
There are usually three types of objective lenses for microscopes used in microbiological research: oil low-magnification objective lens (10×), high-magnification objective lens (40×), and oil lens (100×). There is also the word "OI" (oil immersion) indicating that it has the largest magnification among the three. Depending on the use of eyepieces with different magnifications, the object being inspected can be magnified 1000-1600 times. When used, the difference between oil lenses and other objective lenses is that there is not a layer of air between the slide and the objective lens, but a layer of oil, which is called an oil immersion system. Cedar oil is often used as this oil because the refractive index of cedar oil is n=1.52, which is the same as glass. When light passes through the glass slide, it can enter the objective lens directly through the cedar oil without refraction. If the medium between the glass slide and the objective lens is air, it is called a dry system. When the light passes through the glass slide, it is refracted and scattered, and the light entering the objective lens is obviously reduced, which reduces the illumination of the field of view. The use of oil lenses can not only increase the illumination, but also increase the numerical aperture, because the magnification efficiency of the microscope is determined by its numerical aperture. The so-called numerical aperture is the product of half the sine of the maximum angle at which light is projected onto the objective lens (called the lens angle) multiplied by the refractive index of the medium between the glass plate and the objective lens. It can be expressed by the following formula: NA=n×sinа where NA= Numerical aperture; n = refractive index of the medium; a = half of the maximum incident angle, that is, half of the mirror mouth angle. Therefore, the greater the angle at which light strikes the objective, the greater the efficiency of the microscope. The size of this angle is determined by the diameter and focal length of the objective. At the same time, the theoretical limit of a is 90. . sin90. =1, so when air is used as the medium (n=1), the numerical aperture cannot exceed 1. For example, when cedar oil is used as the medium, n increases, and the numerical aperture also increases. For example, if the incident angle of light is 120o, and its half sine is sin60o=0.87, then: when air is used as the medium: NA=1×0.87=0.87 When water is used as the medium: NA=1.33×0.87=1.15 When cedar oil is used as the medium : NA=1.52×0.87=1.32 The resolution of a microscope refers to the ability of the microscope to distinguish the minimum distance between two points. It is directly proportional to the numerical aperture of the objective lens and inversely proportional to the wavelength length. Therefore, the larger the numerical aperture of the objective lens and the shorter the wavelength of the light wave, the greater the resolution of the microscope and the more clearly the fine structure of the object being inspected can be distinguished. Therefore, a high resolution means a small resolvable distance. These two factors are inversely related. Usually some people talk about the resolution as micrometers or nanometers. This actually confuses the resolution with the minimum resolution distance. Woke up. The resolving power of a microscope is expressed by the minimum distance that can be resolved. The minimum distance that can be distinguished between two points = λ/2NA where λ = wavelength of light wave. The average length of light waves that our naked eyes can feel is 0.55 μm. If a high-power objective lens with a numerical aperture of 0.65 is used, it can distinguish the distance between two points. is 0.42μm. The distance between two points below 0.42 μm cannot be distinguished. Even if a larger eyepiece is used to increase the total magnification of the microscope, it still cannot be distinguished. The only solution is to use a larger objective lens with a larger numerical aperture to increase its resolution. For example, when using an oil lens with a numerical aperture of 1.25, the minimum distance between two points that can be distinguished = 0.55/(2×1.25)=0.22μm. Therefore, we can see that if a high-power objective lens (NA=0.65) with a magnification of 40 times and an eyepiece with a magnification of 24 times are used, although the total magnification is 960 times, the minimum distance of resolution is only 0.42 μm. If you use an oil lens with a magnification of 90 times (NA = 1.25) and an eyepiece with a magnification of 9 times, although the total magnification is 810 times, you can resolve a distance of 0.22 μm.
