Application and use of fluorescence microscope principles
Principles and structural characteristics of fluorescence microscopes: Fluorescence microscopes use a point light source with high luminous efficiency to emit light of a certain wavelength (such as ultraviolet light 3650 in or purple-blue light 4200 in) through a color filter system as excitation light to excite fluorescence in the specimen. After the substance emits fluorescence of various colors, it is then observed through the magnification of the objective lens and eyepiece. In this way, under a strong contrasting background, even if the fluorescence is very weak, it is easy to identify and has high sensitivity. It is mainly used for the study of cell structure and function as well as chemical composition. The basic structure of a fluorescence microscope is composed of an ordinary optical microscope plus some accessories (such as fluorescence light source, excitation filter, two-color beam splitter, blocking filter, etc.). Fluorescent light source - generally uses an ultra-high-pressure mercury lamp (50 to 200W), which can emit light of various wavelengths, but each fluorescent substance has an excitation wavelength that produces the strongest fluorescence, so an excitation filter ( There are generally ultraviolet, purple, blue and green excitation filters), which only allow the excitation light of a certain wavelength to pass through and illuminate the specimen, while absorbing all other light. After each substance is irradiated by excitation light, it emits visible fluorescence with a longer wavelength than the irradiation wavelength in a very short time. Fluorescence is specific and is generally weaker than the excitation light. In order to observe specific fluorescence, a blocking (or suppressing) filter is required behind the objective lens. It has two functions: one is to absorb and block the excitation light from entering the eyepiece to avoid disturbing the fluorescence and damaging the eyes; the other is to select and allow specific fluorescence to pass through to show a specific fluorescence color. The two filters must be selected and used together.
