Following structure is required to take this image in the microscope.
Dark field condensor
Cathode Ray Tube
The image shown is of a Fluorescence Microscopy.
Working Principle of Fluorescence Microscopy.
- Many substances absorb light. However some of them, after absorbing light of a particular wavelength and energy, emit light of a longer wavelength and lesser energy.
- Such substances are called ‘fluorescent substances’.
- Application of this phenomenon is the basis of fluorescence microscope.
- In practice, microbes are stained with a fluorescent dye and then illuminated with blue light. The dye absorbs blue light (shorter wavelength) and emits green light (longer wavelength).
- In a fluorescence microscope, a high intensity mercury arc lamp is used as the light source.
- It emits white light, which is passed through an ‘exciter filter’. It allows only the blue component of the white light to pass through it and blocks out all other color components.
- A dichroic mirror, which reflects blue light, but allows green light is used on the path of the blue light. The mirror is fixed at such an angle that the blue light is reflected downward to the specimen.
- The specimen is previously stained with a fluorescent dye, such as acridine orange NO, acridine yellow,etc. Certain portions of the specimen retain the dye, while others do not. The portions, which retain the fluorescent dye, absorb blue light and emit green light. The emitted green light goes upward and passes through the dichroic mirror.
- Then, the light reaches a ‘barrier filter’. It allows green light to pass to eye .
- Thus, the eye perceives the stained portions of the specimen as glowing green object against a jet-black background, whereas the unstained portions of the specimen remain invisible.