Several Important Optical Technical Parameters of Microscopes
1、 Numerical aperture (N.A.)
Numerical aperture is a key factor in determining the performance of objective lenses (resolution, depth of focus, and brightness).
The numerical aperture (N.A.) is calculated using the following equation.
N. A.=n × Sinx
N=refractive index of the medium between the sample and the objective lens (air: n=1, oil: n=1.515)
X: The angle formed by the optical axis and the refracted light far from the center of the objective lens.
When observing under a microscope, if you want to increase the NA value, the aperture angle cannot be increased. The solution is to increase the refractive index n value of the medium. Based on this principle, water immersed objective lenses and oil immersed objective lenses are produced. Since the refractive index n of the medium is greater than one, the NA value can be greater than one.
The maximum numerical aperture value is 1.4, which has reached its theoretical and technical limit. At present, bromonaphthalene with a high refractive index is used as a medium, and the refractive index of bromonaphthalene is 1.66, so the NA value can be greater than 1.4.
It must be pointed out here that in order to fully utilize the numerical aperture of the objective lens, the NA value of the condenser lens should be equal to or slightly greater than the NA value of the objective lens during observation,
Numerical aperture is closely related to other technical parameters, as it almost determines and affects other technical parameters. It is proportional to resolution, proportional to magnification, and inversely proportional to focal depth. As the NA value increases, the field of view width and working distance will correspondingly decrease.
2、 Resolution
Resolution, also known as "discrimination rate" or "resolution". It is another important technical parameter for measuring the performance of microscopes.
The resolution of the microscope is expressed by the formula: d=l/NA
In the formula, d is the minimum resolution distance; L is the wavelength of the light; NA is the numerical aperture of the objective lens. The resolution of a visible objective is determined by two factors: the NA value of the objective and the wavelength of the illuminating light source. The higher the NA value, the shorter the wavelength of the illumination light, the smaller the d value, and the higher the resolution.
To improve resolution, i.e. reduce the d value, the following measures can be taken
1. Reduce the wavelength l value and use a short wavelength light source.
2. Increase the n value of the medium and increase the NA value (NA=nsinu/2).
3. Increase the aperture angle.
4. Increase the contrast between light and dark.
3、 Magnification rate
Magnification is the magnification, which refers to the ratio of the size of the final image seen by the human eye to the size of the original object after being magnified by the objective lens and then by the eyepiece. It is the product of the magnification of the objective lens and the eyepiece.
Magnification is also an important parameter of a microscope, but one cannot blindly believe that higher magnification is better. When selecting, the numerical aperture of the objective lens should be considered first.
4、 Focal depth
Focal depth is the abbreviation for focal depth, which means that when using a microscope, when the focus is aligned with an object, not only can the points located on the plane of the point be clearly seen, but also within a certain thickness above and below the plane. The thickness of this clear part is called focal depth.
You can see the entire layer of the object being tested, while if the focal depth is small, you can only see a thin layer of the object being tested. The focal depth is related to other technical parameters as follows:
1. The depth of focus is inversely proportional to the total magnification and the numerical aperture of the objective lens.
2. Large depth of focus and reduced resolution.
Due to the large depth of field of the low-power objective, it causes difficulties when taking photos with the low-power objective. A detailed introduction will be provided during micrography. V Field of view diameter
When observing a microscope, the bright prototype range seen is called the field of view, and its size is determined by the field of view aperture in the eyepiece.
The diameter of the field of view, also known as the width of the field of view, refers to the actual range of the object being inspected that can be accommodated within a circular field of view seen under a microscope. The larger the diameter of the field of view, the easier it is to observe.
From the formula, it can be seen that:
1. The diameter of the field of view is proportional to the number of fields of view.
2. Increasing the magnification of the objective lens reduces the field of view diameter. Therefore, if the full view of the object being tested can be seen under a low-power lens, and replaced with a high-power objective, only a small part of the object being tested can be seen.
6、 Working distance
The working distance, also known as object distance, refers to the distance between the surface of the front lens of the objective lens and the object being tested. During microscopic examination, the object being examined should be between one and two times the focal length of the objective lens. Therefore, it and focal length are two concepts, and what is commonly referred to as focusing is actually adjusting the working distance.
When the numerical aperture of the objective lens is fixed, the shorter the working distance, the larger the aperture angle.
A high-power objective with a large numerical aperture has a small working distance.
7、 Poor coverage
The optical system of the microscope also includes a cover glass. Due to the non-standard thickness of the cover glass, the optical path of light entering the air through the cover glass undergoes a change in refraction, resulting in a difference in coverage. The generation of poor coverage affects the sound quality of the microscope.
According to international regulations, the standard thickness of cover glass is 0.17mm,
The allowable range is 0.16-0.18mm, and the difference in this thickness range has been calculated in the manufacturing of the objective lens. The label on the objective lens shell is indeed 0.17, indicating that the thickness of the cover glass required for the objective lens.
