Microscope

There are four basic kinds of microscopes: (1) optical, or light; (2) electron; (3) scanning probe; and (4) ion. This article discusses mainly optical microscopes.

How an optical microscope works. An optical microscope has one or more lenses that refract (bend) the light rays that shine through, or are reflected by, the specimen being observed (see Lens). The refracted light rays make the specimen appear much larger than it is.

The simplest optical microscope is a magnifying glass, which has only one lens (see Magnifying Glass). The best magnifying glasses can magnify an object by 10 to 20 times.

Magnification power is symbolized by a number and the abbreviation X. For example, a 10X magnifying glass magnifies a specimen by 10 times.

A compound microscope uses two or more sets of lenses to provide higher magnifications. Each set of lenses functions as a unit and is referred to as a lens system. A compound microscope has an eyepiece, or ocular, lens system, often called simply the ocular; and one or more objective lens systems, often called objectives.

In microscopes with only one objective, that lens system and the ocular are mounted at opposite ends of a tube. The objective produces a magnified image of the specimen. The ocular then magnifies this image.

In microscopes with two or more objectives, the objectives are mounted in a rotating nosepiece connected to the end of the tube opposite the ocular. The person operating the microscope rotates the nosepiece to align one of the objectives with the opening in the end of the tube. This objective works with the ocular to provide the desired magnification. Many compound microscopes have three objectives that magnify by 4X, 10X, and 40X. When used with a 10X ocular, these microscopes provide magnifications of 40X, 100X, and 400X.

In addition to magnifying a specimen, a microscope must produce a clear image. This capability is called the resolving power, or resolution, of the microscope. In an optical microscope, the wavelength (distance between wave crests) of the light waves that illuminate the specimen limits the resolving power. The wavelength of visible light ranges from about 4,000 to 7,000 angstroms. (One angstrom equals 1/10,000,000 millimeter--about 1/250,000,000 inch). The best optical microscopes cannot resolve parts of a specimen that are closer together than about 2,000 angstroms. To obtain higher resolutions, scientists use other types of microscopes.

Parts of an optical microscope. Most optical microscopes used for teaching have three main parts: (1) the tube, which has already been described; (2) the foot; and (3) the body. The foot is the base of the instrument. The body is an upright support that holds the tube. At the lower end of the body is a mirror. The specimen lies on the stage, a platform above the mirror. The mirror reflects light up through an opening in the stage to illuminate the specimen. The operator can move the tube within the body by turning a coarse-adjustment knob. This focuses the microscope. A fine-adjustment knob moves the tube a small distance for final focusing of a high-power objective.

The diagram above shows the external parts of an optical microscope. A person adjusts these parts to view a specimen. The cutaway diagram at the right shows the path that light follows when passing through the specimen and then through the lenses of the microscope. From The World Book™ Multimedia Encyclopedia ©1998 World Book, Inc., 525 W. Monroe, Chicago, IL 60661. All rights reserved. World Book diagram.

Using an optical microscope. To prepare a microscope for use, turn the nosepiece to bring the objective with the lowest power into viewing position. Next, turn the coarse-adjustment knob to lower the tube until the objective is just above the opening in the stage. Finally, look through the ocular and adjust the mirror so a bright circle of light appears.

Many specimens viewed through a microscope are transparent or have been made transparent. The technique of preparing specimens is called microtomy. Specimens are mounted on glass slides that measure 3 inches long and 1 inch wide (76 by 25 millimeters).

To view a slide, place it on the stage with the specimen directly over the opening. Hold the slide in place with clips that are attached to the stage. Look through the ocular and turn the coarse-adjustment knob to raise the objective until the specimen comes into focus. To avoid breaking the slide or the objective, never lower the lens when a slide is on the stage.

Advanced optical microscopes are used in research and have extra-powerful lens systems. Many such microscopes have a 100X objective and a 20X ocular, which together provide a magnification of 2,000X. Some high-powered microscopes have oil immersion objectives. The bottom lens of the objective touches a drop of special oil placed on the slide. The oil refracts light in a way that provides a better image at high magnification.

Other microscopes used for research have additional features. For example, a microscope with a binocular tube splits the light from the objective into two beams. An ocular for each beam enables the operator to view the specimen with both eyes. Trinocular tubes split the light into three beams--one for each eye and one for a built-in camera. A stereoscope microscope provides a three-dimensional view of the specimen. Such a microscope has an objective and an ocular for each eye.

Scientists use special optical microscopes to study details that are not normally visible. For example, a phase contrast microscope changes the phase relationship between the light waves passing through the specimen and those not passing through it. This action makes some parts of the specimen appear brighter and other parts darker than normal. Parts of a transparent object that vary in thickness or have certain other optical properties can be made visible in this way.

A dark-field microscope prevents light from the lamp from shining directly up the tube. Instead, the microscope uses only light scattered by the specimen. The specimen appears bright against a black background.

A scanning optical microscope does not illuminate the entire specimen at once. Instead, the microscope directs a laser beam at a small spot on the specimen. An electronic device called a photodetector measures the amount of light reflected from, or shining through, the specimen. The beam then scans the specimen as the photodetector takes measurements at a large number of spots. A computer combines the measurements to produce an image on a TV-like monitor screen.

In a confocal scanning optical microscope, a photodetector measures light readings from various depths in the specimen. A computer uses the measurements to create a three-dimensional image. The operator can rotate the image to examine it from any angle.

Other kinds of microscopes. Electron microscopes use a beam of electrons rather than a beam of light to produce magnified images. Electron wavelengths are much shorter than those of visible light. As a result, electron microscopes can resolve much finer detail than optical microscopes can. Some electron microscopes can resolve objects that are less than 2 angstroms apart, which is sufficient to reveal the fundamental atomic structure of the specimen.

There are two basic types of electron microscopes. A transmission electron microscope passes a broad beam of electrons through a specimen slice a few hundred angstroms thick. A scanning electron microscope scans a focused beam across the surface of the specimen. See Electron microscope.

Scanning probe microscopes scan a specimen with a sharp point called a probe. There are two main types of scanning probe microscopes: (1) the scanning tunneling microscope (STM) and (2) the atomic force microscope (AFM). In the STM, the probe does not quite touch the specimen. An electric current flows between the probe and the specimen. A computer uses measurements of this current to create an image. An STM can resolve surface atoms that are less than 2 angstroms apart.

In the AFM, the probe usually gently touches the specimen's surface. As the probe scans the specimen, it reacts to the roughness of the surface by moving up and down. Electronic devices measure this movement and send their results to a computer, which creates an image. The AFM can produce images of specimens, such as animal tissue, through which electric current does not flow readily.

An ion microscope, also known as a field-ion microscope, is used to examine metals. It creates an image of the crystal structure of the tip of an extremely sharp metal needle. An electric field applied to the tip repels charged helium, neon, or argon atoms, which spread out and strike a special screen. The screen glows where the atoms strike it, forming an image of the arrangement of atoms in the metal.

History of the microscope. Engravers probably used water-filled glass globes as magnifying glasses at least 2,000 years ago. The Romans may have made magnifying glasses from rock crystal. Glass lenses of the type now used were introduced in the late 1200's.

Historians generally credit a Dutch spectacle-maker, Zacharias Janssen, with discovering the principle of the compound microscope about 1590. In the 1670's, Anton van Leeuwenhoek, a Dutch amateur scientist, made single-lens microscopes that could magnify up to 270X. Leeuwenhoek was the first person to observe microscopic life and record his observations.

Few improvements occurred until the early 1800's, when better glass-making methods produced lenses that provided undistorted images. German scientists demonstrated the first electron microscope in 1931. The ion microscope was invented in 1951. In 1981, Swiss and West German scientists demonstrated the first scanning tunneling microscope.

Contributor: Stephen J. Pennycook, Ph.D., Senior Research Scientist, Oak Ridge National Laboratory.

Ford, Brian J. The Leeuwenhoek Legacy. Biopress, 1991.

Stewart, Gail B. Microscopes. Lucent Bks., 1992

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