The ability to view things that are too small to be seen by the unaided eye is important in espionage and security. For example, the diagnosis of an infection often relies in part on the visual examination of the microorganism. Information about how the microbe reacts to certain staining methods (e.g, the bacterial Gram stain), the shape of the microbe, and the reaction of antibodies to the microbe all provide important clues as to the identity of the organism.
As well, microscopic examination of documents can reveal information that cannot otherwise be seen. The high magnification and analysis of the elements that make up a sample that is possible using specialized techniques of scanning and transmission electron microscopy can reveal the presence of material that is of suspicious origin (i.e., missile casing), or the presence of codes on a surface.
A microscope is the instrument that produces the highly magnified image of an object that is otherwise difficult or impossible to see with the unaided eye. A microscope is able to distinguish two objects from one another that could not be distinguished with the eye. The resolving power of a microscope is greater than that of the eye.
History of the microscope. In ancient and classical civilizations, people recognized the magnifying power of curved pieces of glass. By the year 1300, these early crude lenses were being used as corrective eyeglasses.
In the seventeenth century Robert Hooke published his observations of the microscopic examination of plant and animal tissues. Using a simple two-lens compound microscope, he was able to discern the cells in a thin section of cork. The most famous microbiologist was Antoni van Leeuwenhoek. Using a single-lens microscope that he designed, Leeuwenhoek described microorganisms in environments such as pond water. His were the first descriptions of bacteria and red blood cells.
By the mid-nineteenth century, refinements in lens grinding techniques had improved the design of light microscopes. Still, advancement was mostly by trial and error, rather than by a deliberate crafting of a specific design of lens. It was Ernst Abbe who first applied physical principles to lens design. Abbe combined glasses that bent light beams to different extents into a single lens, reducing the distortion of the image.
The resolution of the light microscope is limited by the wavelength of visible light. To resolve objects that are closer together, the illuminating wavelength needs to be smaller. The adaptation of electrons for use in microscopes provided the increased resolution.
In the mid-1920s, Louis de Broglie suggested that electrons, as well as other particles, should exhibit wavelike properties similar to light. Experiments on electron beams a few years later confirmed this hypothesis. This was exploited in the 1930s in the development of the electron microscope.
Electron microscopy. There are two types of electron microscope. They are the transmission electron microscope (TEM) and the scanning electron microscope (SEM). The TEM transmits electrons through a sample that has been cut so that it is only a few molecules thin. Indeed, the sample is so thin that the electrons have enough energy to pass right through some regions of the sample. In other regions, where metals that were added to the sample have bound to sample molecules, the electrons either do not pass through as easily, or are restricted from passing through altogether. The different behaviors of the electrons are detected on special film that is positioned on the opposite side of the sample from the electron source.
The combination of the resolving power of the electrons, and the image magnification that can be subsequently obtained in the darkroom during the development of the film, produces a total magnification that can be in the millions.
Because TEM uses slices of a sample, it reveals internal details of a sample. In SEM, the electrons do not penetrate the sample. Rather, the sample is coated with gold, which causes the electrons to bounce off of the surface of the sample. The electron beam is scanned in a back and forth motion parallel to the sample surface. A detector captures the electrons that have bounced off the surface, and the pattern of deflection is used to assemble a three dimensional image of the sample surface.
Scanning, tunneling, and other microscopy techniques. In the early 1980s, the technique called scanning tunneling microscopy (STM) was invented. STM does not use visible light or electrons to produce a magnified image. Instead, a small metal tip is held very close to the surface of a sample and a tiny electric current is measured as the tip passes over the atoms on the surface. When a metal tip is brought close to the sample surface, the electrons that surround the atoms on the surface can actually "tunnel through" the air gap and produce a current through the tip. The current of electrons that tunnels through the air gap is dependent on the width of the gap. Thus, the current will rise and fall as the tip encounters different atoms on the surface. This current is then amplified and fed into a computer to produce a three dimensional image of the atoms on the surface.
Without the need for complicated magnetic lenses and electron beams, the STM is far less complex than the electron microscope. The tiny tunneling current can be simply amplified through electronic circuitry much like that used in other equipment, such as a stereo. In addition, the sample preparation is usually less tedious. Many samples can be imaged in air with essentially no preparation. For more sensitive samples that react with air, imaging is done in vacuum. A requirement for the STM is that the samples be electrically conductive.
Scanning tunneling microscopes can be used as tools to physically manipulate atoms on a surface. This holds out the possibility that specific areas of a sample surface can be changed.
Other forces have been adapted for use as magnifying sources. These include acoustic microscopy, which involves the reflection of sound waves off a specimen; xray microscopy, which involves the transmission of x rays through the specimen; near field optical microscopy, which involves shining light through an opening smaller than the wavelength of light; and atomic force microscopy, which is similar to scanning tunneling microscopy but can be applied to materials that are not electrically conductive, such as quartz.
█ FURTHER READING:
Aebi, Engel. Atlas of Microscopy Techniques. San Diego: Plenum Press, 2002.
Hayat, M. Arif. Microscopy, Immunohistochemistry, and Antigen Retrieval Methods for Light and Electron Microscopy. New York: Plenum Publishing, 2002.
Murphy, Douglas, B. Fundamentals of Light Microscopy and Electronic Imaging. New York: Wiley-Liss, 2001.