Physics: Microscopy

Introduction

Though not a scientific field per se, microscopy (the use of one of many types of microscopes) is an important technological tool within many scientific disciplines. By allowing scientists to view samples of various materials at magnification, microscopy exposes details that were previously invisible. The use of microscopes has led to the discovery of the cell and the organelles within it, both crucial understandings in the field of biology, as well as many other discoveries.

Since its invention about 400 years ago, the microscope has undergone many changes and improvements, especially in professional models. Many of the most powerful microscopes no longer use visible light to form their images but employ electrons to create high-resolution images of very tiny objects, enabling researchers to study their structure. On the other hand, student-model microscopes have changed very little since their invention. Their continued use in education shows the power of this relatively simple tool.

Historical Background and Scientific Foundations

The most fundamental part of a microscope is a magnifying lens. Though rare, lenses of this type were produced in ancient times. During excavations in Nineveh, a city in ancient Mesopotamia, a quartz-crystal lens dating from around 640 BC was unearthed. Similar lenses have been found on the island of Crete and other ancient Greek sites. Some of these lenses could provide magnification up to 20 times, though most were only useful for much lesser magnification. The Greeks also used “burning glasses” to concentrate the sun's rays, and both Greeks and Romans used glass spheres filled with water as magnifiers. Engravers probably used lenses to create fine carvings, and philosophers such as Pliny the Elder (AD 23–79) were interested in them as objects of scientific curiosity.

During the Renaissance significant advances were made in the field of optics, when technology became reliable enough to make lenses that corrected vision. One of the most famous scientists who worked with optics was Italian physicist, astronomer, and mathematician Galileo Galilei (1564–1642). Galileo is especially known for his telescopes, which he built and used to study the solar system. His observations of Jupiter led to the discovery of several of its moons. Galileo also made an early compound microscope that he shared with his colleagues in the Academia dei Lincei, one of the earliest scientific academies.

Some confusion exists about the date of the microscope's invention and its inventor. Many credit Dutch opticians Hans Jansen and his son Zacharias (1585–1632), placing their invention around 1600; others cite another Dutch spectacle maker, Hans Lippershey (c.1570–c.1619). The Jansens' machine was a compound microscope, with a tube connecting a lens at the eyepiece to another near the sample. Both Galileo's device and the Jansens' were based on the design of telescopes, another early object of scientific curiosity.

The microscope allowed many kinds of scientists to study their fields in more depth than ever before. One of these was Marcello Malpighi (1628–1694), an Italian physiologist and one of the first to study microscopic anatomy. Malpighi questioned the fundamental principals of medicine in his day. He used his microscopic observations to prove, among other things, that blood was continuously circulated, not turned to flesh when it reached the edges of the body. He also studied the tissues of the brain, liver, kidneys, and other organs, giving the first descriptions of their microscopic features.

Today, Malpighi is considered to be a key pioneer of histology, the study of tissues, and is also very important to early work in embryology.

One of the most influential early microscopists was Robert Hooke (1635–1703), an English scientist and mathematician who made important contributions in many scientific fields. Hooke modified the Jansen compound microscope to suit his own uses and to make it more powerful. He also devised a system to illuminate his subjects for better viewing. By slicing cork very thinly, he discovered and named the plant cell. Hooke also made observations on many other kinds of plants and animals, publishing them in his seminal book called Micrographia (Small drawings) in 1665. It contained his observations on many different organisms, including insects, sponges, and bird feathers. Hooke also used his position as curator of experiments for the Royal Society of London to demonstrate his microscopic findings before many important scientists of the day.

Another giant of microscopy was Antonie van Leeuwenhoek (1632–1723), an uneducated Dutch cloth trader. Descended from basket makers and brewers, van Leeuwenhoek seems an unlikely man to revolutionize the science of biology. After studying Hooke's Mi-crographia, van Leeuwenhoek became fascinated with microscopy. He was familiar with magnifiers from his work in the cloth trade, where they were used to examine goods. He decided to build his own microscope, which he manufactured himself. Instead of using Hooke's popular compound design, van Leeuwenhoek constructed what amounted to extremely powerful magnifying glasses with powers of up to 200 times. His simple devices were easy and quick to make, though not always easy to use, and he made up to 500 of them during his life.

Using his magnifiers, van Leeuwenhoek began to explore the world around him on a microscopic level. His investigations of common substances such as pond water, tooth plaque, blood, semen, animal tissue, plant sections, minerals, crystals, and fossils led to his many discoveries. In the course of his investigations, van Leeu-wenhoek discovered bacteria, protists, the algae Spirogyra, blood cells, sperm cells, and microscopic worms. He became a member of the Royal Society of London, and, though he never visited it, his correspondence laid

MARCELLO MALPIGHI (1628–1694)

In the second half of the seventeenth century, Italian physician Marcello Malpighi (1628–1694) used the newly invented microscope to make a number of important discoveries about living tissues and structures, earning himself enduring recognition as a founder of scientific microscopy, histology (the study of tissues), embryology, and the science of plant anatomy.

Early in his medical career, Malpighi became absorbed in using the microscope to study a wide range of living tissue—animal, insect, and plant. At the time, this was an entirely new field of scientific investigation. Malpighi soon made a profoundly important discovery. Microscopically examining a frog's lungs, he was able for the first time to describe the lung's structure accurately—thin air sacs surrounded by a network of tiny blood vessels. This explained how air (oxygen) is able to diffuse into the blood vessels, a key to understanding the process of respiration. It also provided the one missing piece of evidence to confirm William Harvey's (1578–1657) revolutionary theory of blood circulation: Malpighi had discovered the capillaries, the microscopic connecting link between the veins and arteries that Harvey—with no microscope available—had only been able to postulate. Malpighi published his findings about the lungs in 1661.

Malpighi used the microscope to make an impressive number of other important observations, all “firsts.” He observed a “host of red atoms” in the blood—the red blood corpuscles. He described the papillae of the tongue and skin—the receptors of the senses of taste and touch. He identified the rete mucosum, the Malpighian layer, of the skin. He found that the nerves and spinal column both consisted of bundles of fibers. He clearly described the structure of the kidney and suggested its function as a urine producer. He identified the spleen as an organ, not a gland; structures in both the kidney and spleen are named after him. He demonstrated that bile is secreted in the liver, not the gall bladder. In showing bile to be a uniform color, he disproved a 2,000-year-old idea that bile was yellow and black. He described glandular adenopathy, a syndrome rediscovered by Thomas Hodgkin (1798–1866) and given that man's name 200 years later.

Malpighi also conducted groundbreaking research in plant and insect microscopy. His extensive studies of the silkworm were the first full examination of insect structure. His detailed observations of chick embryos laid the foundation for microscopic embryology. His botanical investigations established the science of plant anatomy. The variety of Malpighi's microscopic discoveries piqued the interest of countless other researchers and firmly established microscopy as a science.

out many of his significant discoveries. Amazingly, in 1981 several of van Leeuwenhoek's original samples were discovered in the Royal Society's archives by mi-croscopist Brian J. Ford (1939–), whose reanalysis of the original specimens revealed the excellent quality of van Leeuwenhoek's work and once again confirmed its importance to science as a whole.

With improved lenses and mechanical fittings, the compound microscope became the most powerful and widely used magnifying instrument. The development of the stage and slides that rest upon it permitted better examination of samples. Staining also improved visibility. Today microscopy remains a useful tool within many scientific and technical fields. Three major types of microscopy are used to examine increasingly minute samples, providing more information than ever before.

The most recognizable type of microscope is the optical compound microscope. It developed directly from the earliest compound microscopes and shares many of the same features. The most significant development for this type of microscope was the addition of an artificial light source, which allowed consistent observation of samples. Different light sources can be used at different wavelengths or from various angles to illuminate indistinct structures. New ways of viewing optical microscope images have also been developed. Stereomicroscopes create two images from slightly different angles and transmit them through two eyepieces, producing a three-dimensional view of the subject. Cameras can also be used to view the sample, creating photographic or digital images.

Electron microscopes use electrons, not light, to visualize samples, permitting much greater magnification. There are two major types. Transmission electron microscopes (TEM) work in a way similar to optical microscopes. They pass a beam of electrons through a sample and use a detector to form the image. The sample must be sliced into an extremely thin section for the beam to pass through it. Advanced-transmission electron microscopes have been able to deliver magnification of up to 50 million times, allowing even individual atoms to be visualized. This gives it an important role in nano-technology development. The other type of electron microscope, the scanning electron microscope (SEM), is responsible for many dramatic and captivating images, as well as important discoveries. Its electron beam scans a sample's surface, producing an image with great depth of field (large portions of the image are in focus) giving it an almost three-dimensional appearance.

Transmission electron microscopes (TEM) and scanning electron microscopes (SEM) offer important

variations on basic electron microscopy. The TEM transmits electrons through an extremely thin sample. The electrons scatter as they collide with the atoms in the sample and form an image on a photographic film below the sample. This process is similar to a medical x ray, where x rays (very short wavelength light) are transmitted through the body and form an image on photographic film behind the body.

By contrast, the SEM reflects a narrow beam of electrons off the surface of a sample and detects the reflected electrons. To image a certain area of the sample, the electron beam is scanned in a back and forth motion parallel to the sample surface, similar to the process of mowing a square section of lawn.

The chief differences between the two microscopes are that the TEM gives a two-dimensional picture of the interior of the sample while the SEM gives a three-dimensional picture of the surface of the sample. Images produced by SEM are familiar to the public, as in television commercials showing pollen grains or dust mites.

In the early 1980s, a new technique in microscopy was developed which did not involve beams of electrons or light to produce an image. Instead, a small metal tip is scanned 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. Some probes are so sharp that the tip is composed of a single atom. The first scanning probe microscope, called a scanning tunneling microscope (STM), used a quantum property of electrons to examine a sample. Now many different types are specialized to gather different types of data, not just produce images, although they can be used to do so. Scanning probe devices are not exactly microscopes—the image is created not by “looking” at the surface, but by “feeling” it with the probe. 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. This physical phenomenon is called tunneling and is one of the amazing results of quantum physics. If such a phenomenon could occur with large objects, it would be possible for abaseball to tunnel through a brick wall with no damage to either. The current of electrons that tunnels through the air gap is very much dependent on the width of the gap; therefore the current will rise and fall in succession with the 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 similar to circuitry that is used in other electronic 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 conducting, such as a metal.

Modern Cultural Connections

Some of the first scientific investigations using microscopes were in the fields of biology and the life sciences. Hooke and van Leeuwenhoek's discoveries have led to even more important advances within life sciences. Hooke's discovery of the cell was made possible by the microscope, as was the later discovery of the organelles within it. Observing these structures and learning their functions have revealed the inner workings of the cell: metabolism and reproduction. Through the use of the microscope, a vast number of different cell and tissue types have been identified. By understanding their structures and roles within the body, scientists have learned more about the way life functions.

In zoology the microscope has been invaluable for studying both extremely small organisms and the tiny features present on creatures of all sizes. In Micrographia, Hooke observed a flea and drew pictures of it in minute detail. Insects are often studied by microscopy today, including some species that may be too small to study otherwise. Insects' tiny scales and hairs, complicated eye structure, and different body plans have been revealed by microscopy. Microscopy also revealed the hollow structure of polar bears' hair shafts, which provide extra insulation in their northern habitats. Botanists use microscopes to study plants in many of the same ways zoologists study animals. Both microscopic plants and tiny features of larger ones can be seen with a microscope.

The entire field of microbiology has been made possible by the use microscopes, which have been used to observe bacteria, viruses, fungi, protists, and algae; all are visible only with the aid of a microscope. Studying microorganisms has led to cures for disease, improved

industrial products, and new ways to decompose dangerous waste. The field still holds much promise, as biologists project that only a tiny percentage of existing microorganisms has been discovered and studied. Doctors and medical technicians use microscopes to study many of the same organisms and tissues as other scientists, hoping to cure disease.

Microscopy has become important in many technological fields as well. The minute analysis of materials has led to stronger, lighter, and more durable products. The production of microchips would not be possible without the ability to analyze and build them on a minute scale. The most powerful scanning probe microscopes can already image single atoms, allowing continuing advances in nanotechnology.

The microscope's lasting impact is its ability to reveal features that were previously hidden by their miniscule size. Studying these features has produced a better understanding of the basic function of things as diverse as bacteria and ball bearings. By exploring the tiniest, most basic elements of the world around us, we can better understand its function as a whole.

bibliography

Periodicals

Sines, George, and Yannis A. Sakellarakis. “Lenses in Antiquity.” American Journal of Archaeology 91 (1987): 191–196.

Web Sites

Florida State University. Molecular Expressions: Exploring the World of Optics and Microscopy. “Introduction to Microscopy.” March 15, 2005.http://micro.magnet.fsu.edu/primer/index.html (accessed February 2, 2008).

Microscopy—UK. An Introduction to Microscopy.“The History of the Microscope.” http://www.microscopy-uk.org.uk/index.html?http://www.microscopy-uk.org.uk/intro/histo.html (accessed February 2, 2008).

University of Dayton. “The History of the Microscope.” http://campus.udayton.edu/̃hume/Microscope/microscope.htm (accessed February 2, 2008).

University of Nebraska—Lincoln. Electron Microscopy.“What Are Electron Microscopes?” http://www.unl.edu/CMRAcfem/em.htm (accessed February 2, 2008).

Kenneth T. LaPensee