How can the resolution of a microscope be improved?
The microscope is one of the main equipment for testing equipment, and an important indicator for judging the performance of a microscope is resolution. Resolution refers to the ability to clearly distinguish the smaller distance between two small points or two lines. The human eye itself is a microscope. Under standard lighting conditions, the resolution of the human eye at the photopic distance (internationally recognized as 25cm) is approximately equal to 1/10mm. For observing two straight lines, since straight lines can stimulate a series of nerve cells, the resolution of the eyes can be improved.
The resolution of the human eye is only 1/10mm, so the human eye cannot distinguish objects smaller than 1/10mm or the distance between two tiny objects closer than 1/10mm. So there were simple macroscopic magnifying glasses, optical microscopes for microscopic observation, and then electron microscopes. The resolution of a microscope is defined as the smaller distance between two small points on a specimen that can be clearly distinguished. The calculation formula is: D=0.61λ/NA
In the formula: D is the resolution (um); λ is the wavelength of the light source (um); NA is the numerical aperture of the objective lens (also called the lens aperture ratio).
It can be obtained from the formula that the resolution of the microscope is determined by the wavelength of the incident light source and the numerical aperture of the matching objective lens. It can be seen from this that methods to improve optical microscopy:
1. Reduce the wavelength of the light source.
The shorter wavelength of visible light is 390nm. If ultraviolet light of this wavelength is used as the illumination source, the resolution of the optical microscope can be reduced to 0.2um. However, since most ordinary glass materials absorb a large amount of light with wavelengths below 340nm, ultraviolet light cannot form a clear and bright image after a large amount of attenuation. Therefore, expensive materials such as quartz (which can transmit ultraviolet light as low as 200nm) and fluorite (which can transmit ultraviolet light as low as 185nm) have to be used, and ultraviolet light microscopes cannot be observed with the naked eye, even under the observation of samples. Coupled with the high cost, this method of improving microscope resolution is not widely used due to its own limitations.
2. Increase the numerical aperture NA of the objective lens.
Numerical aperture NA=n*sin(u)
In the formula, n is the refractive index of the medium between the objective lens and the specimen; u is one-half aperture angle of the objective lens. Therefore, appropriately adopting a larger aperture angle in optical design or increasing the refractive index has become a common method to improve the resolution of optical microscopes. Generally, the medium of low-magnification objective lenses such as those below 10X is air, with a refractive index of 1, that is, a dry system objective lens; the water immersion medium is distilled water, with a refractive index of 1.33; the medium of an oil immersion objective lens is cedar oil or other transparent oil, with an average refractive index. At around 1.52, it is close to the refractive index of lenses and glass slides, such as Olympus' 100X oil lens. Water immersion objectives and oil immersion objectives not only have high magnification, but also use high refractive index media to improve the resolution of the objective lens.
