Fluorescence microscopes can be divided into two types according to the principle of light path:
1. Transmission fluorescence microscope
In older fluorescence microscopes, the excitation light source passes through the specimen material through a condenser to excite fluorescence. The advantage is that the fluorescence is strong at low magnification, but the disadvantage is that the fluorescence weakens as the magnification increases. So it is only suitable for observing larger specimen materials.
2.Epifluorescence microscopy
The excitation light falls downward from the objective lens onto the specimen surface, i.e., the same objective lens is used as the illumination condenser and the objective lens for collecting fluorescence.
A dichroic beam splitter (dichroic mirror) needs to be added to the optical path, which is at an angle of 45o to the optical axis. The excitation light is reflected into the objective lens and concentrated on the sample. The fluorescence generated by the sample is reflected by the surface of the objective lens and the cover. The excitation light reflected on the surface of the glass slide enters the objective lens at the same time and returns to the dichroic beam splitter to separate the excitation light and fluorescence. The remaining excitation light is then absorbed by the blocking filter. If you use different excitation filter/dual-color beam splitter/blocking filter combination inserts, you can meet the needs of different fluorescence reaction products.
The advantage of this kind of fluorescence microscope is that the field of view is uniformly illuminated, the image is clear, and the greater the magnification, the stronger the fluorescence.
What are the main categories of optical microscopes?
1. Ordinary optical microscope
Ordinary optical microscopes are mainly composed of the following parts: the illumination system, which involves the light source and condenser; the optical magnification system, which is composed of the objective lens and the eyepiece. It is the most critical part of the microscope and is designed to To avoid excessive spherical aberration and chromatic aberration, both the eyepiece and objective lens are composed of complex lens groups.
2. Laser confocal scanning microscope
The laser confocal scanning microscope sounds very high-end and complicated. In fact, it simply uses laser as a scanning light source to quickly scan and image objects point by point, line by line, and surface by surface.
Based on the shorter wavelength of the laser beam, the beam itself is very thin, which determines that the confocal laser scanning microscope has a high resolution, which is about three times that of an ordinary optical microscope. This type of microscope is used to observe cell morphology and quantitatively analyze intracellular biochemical components and measure cell morphology.
3. Dark field microscope
There is a light plate in the center of the condenser of a dark-field microscope, which prevents illumination light from entering the objective lens directly. Only the light reflected and diffracted by the specimen can enter the objective lens. Therefore, the background of the field of view is black and the edges of the objects are bright.
