Principles Of Microscopy

Microscopes make small objects appear bigger. A light microscope will magnify an image up to 1500 times its original size. Electron microscopes can achieve magnifications up to 1 million times. However, bigger is only better when more details are revealed. The fineness of detail that a microscope can reveal is its resolving power. This is defined as the smallest distance that two objects can approach one another yet still be recognized as being separate. The resolution that a microscope achieves is mainly a function of the wavelength of the illumination source it employs. The smaller the wavelength, the smaller the object that will cause diffraction, and the better the resolving power. The light microscope, because it uses visible light of wavelength around 500 nanometers (nm, where 1000 nm = 1 ^.m), can distinguish objects as small as about half this: 250 nm. It can therefore be used to visualize the smallest cells and the major intracellular structures or organelles. The microscopic study of cell structure organization is known as cytology. An electron microscope is required to reveal the ultrastructure (the fine detail) of the organelles and other cytoplasmic structures (Fig. 1.2).

The wavelength of an electron beam is about 100,000 times less than that of white light. In theory, this should lead to a corresponding increase in resolution. In practice, the

Figure 1.2. Cell structure as seen through the light and transmission electron microscopes.

electron microscope can distinguish structures about 1000 times smaller than is possible in the light microscope, that is, down to about 0.2 nm in size.

The Light Microscope

A light microscope (Figs. 1. 3a and 1.4) consists of a light source, which may be the sun or an artificial light, plus three glass lenses: a condenser lens to focus light on the specimen, an objective lens to form the magnified image, and a projector lens, usually called the eyepiece, to convey the magnified image to the eye. Depending on the focal length of the various lenses and their arrangement, a given magnification is achieved. In bright-field

light

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