The microscopes which are based on visible light as a source of illumination have their own limitations. In spite of the numerous modifications there is a limit to the resolving power of these microscopes.
The maximum resolution that the light microscopes can achieve is the wave length of the light waves. A particle smaller than the wavelength of visible light cannot be resolved under the light microscope.
The development of electron microscope in the late 1940’s however has changed the situation dramatically.
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Using a source of electrons for illumination which have a much shorter wave length than that of light rays, the electron microscope has opened up before biologists a hitherto unknown exquisite sub cellular architecture of the world of microcosm.
The electron microscope is a remarkable research tool of the 20th century. Its power of magnification runs into 100000X or a resolving power of 10 centimeters.
Experiments that eventually lead to the development of electron microscope began as early as 1920. The First prototype model was developed for physical applications.
Principle of working:
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As the name indicates, in an electron microscope, a beam of electrons is used as a source of illumination, instead of a beam of light as in light microscopes. In order to understand the working mechanism of an electron microscope
We must first understand some elementary facts about electrons and their behavior. Electrons are negatively charged subatomic particles having a mass of 9.1 x 1028 grams. An integral part of all atoms, electrons orbit round the atomic nucleus at velocities for exceeding 50,000 kms per second.
When the atoms of a metal are excited by heat energy to an extent where the electron velocity exceeds the attraction of the nucleus, they (electrons) fly off from the clutches of the nucleus and are lost.
When a metal, like tungsten is heated by applying a high voltage current, electrons come out in a continuous stream and this can be directed to form a high velocity electron beam.
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Louis de Bragile (1924) working on electrons opined that electrons being particulate must behave like light waves and should have both a fixed wavelength and frequency.
He used the following formula to calculate the wave length X (lambda) of the matter where he is plank’s constant, m is mass and v is velocity. Calculations based on this equation have shown that electrons accelerated to a potential of 60,000 volts have a wave length of 0.05 A (Angstrom) or 105 times smaller than that of visible light.
Hence an electron beam should be able to resolve (theoretically) particles 10 5 times smaller than that of the wave length of light the discovery that electrons have wave motion and they can be focused on to an object in the form of a beam (like light ray) in a magnetic field provided the impetus for the development of electron microscope.
By the late 1930’s a system of magnetic coils capable of focusing an electron beam had been designed it was also shown that by regulating the current flowing through the magnetic coils, the magnification can be regulated.
Theoretically, an enhancement of resolution power equivalent to the wave length of a beam of electrons should be possible in an electron microscope. But there are practical limitations in the design of magnetic coils (equivalent to lenses in a light microscope) used for focusing the beam of electrons.
The main problem is the one of the distortion produced when the angle of illumination is more than a few tenths of a degree as a result; to catch the beam of electrons, the numerical aperture of objectives has to be much lesser than that of light microscopes.
The actual resolving power of electron microscopes is only about 0.05nm; far less than the theoretical potential of 0.002mm but even then, 0.05nm is much smaller when compared with 100 mm that is the limit in a light microscope.