Micro Bio

Early microbiology Ancient cultures and civilizations had no idea that microbes existed but they did comprehend some of their important effects. For example: • • • •

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Ancient Egyptians were among the earliest peoples to use fermentation to brew their own beer. The Romans liked to have good sanitation and prized clean drinking water. Ancient Chinese immunized people against smallpox by having them inhale dried, powdered scabs from those suffering from a mild form of the disease. Many traditional cultures have also recognized and used plants as remedies for certain diseases. For example, South Americans recognized the usefulness of extracts Cinchona tree (containing quinine) to treat malaria. The importance of traditional healers is being rediscovered by the West and has contributed to the field of ethnobotany. Many cultures recognized the communicability of certain diseases. Unfortunately, this recognition led to fear of and discrimination against sick people. These fears still persist today.

A Brief History of Microbiology Development of microscopy: •

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Aristotle (384-322) and others believed that living organisms could develop from non-living materials. 1590: Hans and Zacharias Janssen (Dutch lens grinders) mounted two lenses in a tube to produce the first compound microscope. 1660: Robert Hooke (1635-1703) published "Micrographia", containing drawings and detailed observations of biological materials made with the best compound microscope and illumination system of the time. 1676: Anton van Leeuwenhoek (1632-1723) was the first person to observe microorganisms. 1883: Carl Zeiss and Ernst Abbe pioneered developments in microscopy (such as immersion lenses and apochromatic lenses which reduce chromatic aberration) which perist until the present day. 1931: Ernst Ruska constructed the first electron microscope.

Spontaneous generation controversy: •

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1688: Francesco Redi (1626-1678) was an Italian physician who refuted the idea of spontaneous generation by showing that rotting meat carefully kept from flies will not spontaneously produce maggots. 1836: Theodor Schwann (1810-1882) helped develop the cell theory of living organisms, namely that that all living organisms are composed of one or more cells and that the cell is the basic functional unit of living organisms. 1861: Louis Pasteur's (1822-1895) famous experiments with swan-necked flasks finally proved that microorganisms do not arise by spontaneous generation.

This eventually led to: • •

Development of sterilization Development of aseptic technique

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Proof that microbes cause disease: 1546: Hieronymus Fracastorius (Girolamo Fracastoro) wrote "On Contagion" ("De contagione et contagiosis morbis et curatione"), the the first known discussion of the phenomenon of contagious infection. 1835 Agostino Bassi de Lodi showed that a disease affecting silkworms was caused by a fungus - the first microorganism to be recognized as a contagious agent of animal disease. 1847: Ignaz Semmelweiss (1818-1865), a Hungarian physician who decided that doctors in Vienna hospitals were spreading childbed fever while delivering babies. He started forcing doctors under his supervision to wash their hands before touching patients. 1857: Louis Pasteur proposed the "germ theory" of disease. 1867: Joseph Lister (1827-1912) introduced antiseptics in surgery. By spraying carbolic acid on surgical instruments, wounds and dressings, he reduced surgical mortality due to bacterial infection considerably. 1876: Robert Koch (1843-1910). German bacteriologist was the first to cultivate anthrax bacteria outside the body using blood serum at body temperature. Building on pasteur's "germ theory", he subsequently published "Koch's postulates" (1884), the critical test for the involvement of a microorganism in a disease: 1. The agent must be present in every case of the disease. 2. The agent must be isolated and cultured in vitro. 3. The disease must be reproduced when a pure culture of the agent is inoculated into a susceptible host. 4. The agent must be recoverable from the experimentally-infected host. This eventually led to: • •

Development of pure culture techniques Stains, agar, culture media, petri dishes

Bacteria were first observed by Antonie van Leeuwenhoek in 1676, using a single-lens microscope of his own design.[10] He called them "animalcules" and published his observations in a series of letters to the Royal Society.[11][12][13] The name bacterium was introduced much later, by Christian Gottfried Ehrenberg in 1838.[14] Louis Pasteur demonstrated in 1859 that the fermentation process is caused by the growth of microorganisms, and that this growth is not due to spontaneous generation. (Yeasts and molds, commonly associated with fermentation, are not bacteria, but rather fungi.) Along with his contemporary, Robert Koch, Pasteur was an early advocate of the germ theory of disease.[15] Robert Koch was a pioneer in medical microbiology and worked on cholera, anthrax and tuberculosis. In his research into tuberculosis, Koch finally proved the germ theory, for which he was awarded a Nobel Prize in 1905.[16] In Koch's postulates, he set out criteria to test if an organism is the cause of a disease; these postulates are still used today.[17] Though it was known in the nineteenth century that bacteria are the cause of many diseases, no effective antibacterial treatments were available.[18] In 1910, Paul Ehrlich developed the first antibiotic, by changing dyes that selectively stained Treponema pallidum—the spirochaete that causes syphilis—into compounds that selectively killed the pathogen.[19] Ehrlich had been awarded a 1908 Nobel Prize for his work on immunology, and pioneered the use of stains to detect and identify bacteria, with his work being the basis of the Gram stain and the Ziehl-Neelsen stain.[20] A major step forward in the study of bacteria was the recognition in 1977 by Carl Woese that archaea have a separate line of evolutionary descent from bacteria.[21] This new phylogenetic taxonomy was based on the sequencing of 16S ribosomal RNA, and divided prokaryotes into two evolutionary domains, as part of the three-domain system.[22]

What is a virus? Viruses are very tiny, simple organisms. In fact, they are so tiny that they can only be seen with a special, very powerful microscope called an "electron microscope," and they are so simple that they are technically not even considered "alive." There are six characteristics of all living things:

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Adaptation to the environment Cellular makeup Metabolic processes that obtain and use energy Movement response to the environment Growth and development Reproduction

A virus is not able to metabolize, grow, or reproduce on its own, but must take over a host cell that provides these functions; therefore a virus is not considered "living." The structure of a virus is extremely simple and is not sufficient for an independent life. Structure: Each virus is made up of two elementary components. The first is a strand of genetic material, either deoxyribonucleic acid (DNA) or ribonucleic acid (RNA). Unlike living cells, viruses will have either DNA or RNA, but not both. The genetic material is a blueprint for determining the structure and behavior of a cell. In a virus, a protein coat called a "capsid" surrounds the nucleic acid. This coat serves to protect the nucleic acid and aid in its transmission between host cells. The capsid is made of many small protein particles called "capsomeres," and can be formed in three general shapes – helical, icosahedral (a 20-sided figure with equilateral triangles on each side), and complex. Some of the more advanced viruses have a third structure that surrounds the capsid. This is called the "envelope" and is composed of a bilipid layer, like the membrane on a cell, and glycoproteins, which are protein and carbohydrate compounds. The envelope serves to disguise the virus to look like a 'real' cell, protecting it from appearing as a foreign substance to the immune system of the host. The structure of a virus is closely related to its mode of reproduction. Reproduction: A virus's sole purpose is to reproduce, but it needs a host cell to do so. Once a suitable host cell has been located, the virus attaches to the surface of the cell or is ingested into the cell by a process called "phagocytosis." It then releases its genetic material into the cell, and essentially shuts down normal cell processes. The cell stops producing the proteins that it usually makes and uses the new blueprint provided by the virus to begin making viral proteins. The virus uses the cell's energy and materials to produce the nucleic acid and capsomeres to make numerous copies of the original virus. Once these 'clones' are assembled, the virus causes the host cell to rupture, releasing the viruses to infect neighboring cells.

Viral Shapes Name

Basic Shape

Example (electron micrograph)

Helical

Tobacco mosaic virus

Icosahedral

Herpes simplex

Complex

T-4 Bacteriophage

Hosts and resistance: Viruses are known to infect almost any kind of host that has living cells. Animals, plants, fungi, and bacteria are all subject to viral infection. But viruses tend to be somewhat particular about what type of cells they infect. Plant viruses are not equipped to infect animal cells, for example, though a certain plant virus could infect a number of related plants. Sometimes, a virus may infect one creature and do no harm, but cause havoc when it gets into a different but closely related creature. For example, deer mice carry the Hantavirus without much noticeable effect on the rodents, but if Hantavirus infects a person, it causes a dramatic, and frequently deadly, disease marked by excessive bleeding. Most animal viruses, however, are species specific. This means, they will infect one species of animal. For instance, feline immunodeficiency virus (FIV) will only infect cats; human immunodeficiency virus (HIV) will only infect humans. What does the host do in order to fight off the invasion of a virus? Any foreign substance introduced into the body produces what is called an "immune response." Through this process, the host's body produces antibodies. Antibodies are substances that will destroy an invader and prevent the host from contracting the same disease again in the future. Antibodies are specific to each invader, and each time that a new disease is contracted, a new set of antibodies will have to be manufactured. This process of making antibodies specific to the infecting virus takes about seven days. In the meantime, the cell infected with a virus produces small proteins called "interferons." These interferons are released within three to five days and function to prevent infection in neighboring cells until the antibodies can be made. Needless to say, research on the benefit of interferons in viral treatment is underway, but the actual mechanism of an interferon is not entirely known. There are some antiviral medications that can be administered in the case of viral infection, but the body's immune system is largely relied upon to fight off these kinds of infections.

What are bacteria? Bacteria are very different from viruses. First of all, bacteria are much larger in size. The largest virus is only as big as the very smallest bacterium (singular for bacteria). But bacteria are still microscopic and cannot be seen with the naked eye. They are so small that the sizes of bacteria are measured in micrometers (10,000 micrometers = 1 centimeter). By comparison, the head of a pin is about 1000 micrometers wide. Though more complex than a virus, the structure of a bacterium is still relatively simple. Structure: Most bacteria have an outer, rigid cell wall. This provides shape and support. Lining the inside of the cell wall is a plasma membrane. This is like the membrane found around all living cells that provides both a boundary for the contents of the cell and a barrier to substances entering and leaving. The content inside the cell is called "cytoplasm." Suspended in the cytoplasm are ribosomes (for protein synthesis), the nucleoid (concentrated genetic material), and plasmids (small, circular pieces of DNA, some of which carry genes that control resistance to various drugs). All living cells have ribosomes, but those of bacteria are smaller than those found in any other cell. Some antibacterial medicines have been made that attack the ribosomes of a bacterium, leaving it unable to produce proteins, and therefore killing it. Because the ribosomes are different, the cells of the host are left unharmed by the antibiotic. Other antibiotics target certain portions of the cell wall. Some bacteria have long, whip-like structures called "flagella" that they use for movement. Bacteria can occur in three basic shapes:

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Coccus (spheres) Bacillus (rods) Spirillum (spirals)

Bacterial Shapes Name

Basic Shape

Example (electron micrograph)

Coccus (sphere)

Staphylococcus aureus

Bacillus (rod)

(starting to divide)

Salmonella typhi

Spirillum (spiral)

Campylobacter jejuni

Reproduction: Bacteria undergo a type of asexual reproduction known as "binary fission." This simply means they divide in two, and each new bacterium is a clone of the original – they each contain a copy of the same DNA. Bacteria can reproduce very quickly. In fact, in an ideal laboratory situation, an entire population of bacteria can double in only twenty minutes. At this enormous growth rate, one bacterium could become a BILLION (1,000,000,000) bacteria in just 10 hours! Luckily, there are neither enough nutrients nor space available to support this rapid growth, or the world would be overrun with bacteria. As it is, bacteria can be found living on almost any surface and in almost any climate in the world. Hosts and resistance: As stated, bacteria can grow nearly everywhere. These microbes have been around for billions of years because they are able to adapt to the ever-changing environment. They can find a home anywhere, and some of them live in places where it was once thought 'nothing' could survive. There are bacteria in the soil, at the depths of the ocean, living in the mouth of volcanoes, on the surfaces of teeth, and in the digestive tracts of humans and animals. They are everywhere and are very numerous. For example, a single teaspoon of soil is said to contain at least 1,000,000,000 bacteria. Most often, bacteria are thought of as a bad thing, but most bacteria are not pathogenic (disease-causing). In fact, many bacteria are very helpful to us. There are species that decompose trash, clean up oil spills, and even produce medicines. The few species that are pathogenic, however, give the rest of the bacteria a bad name. Pathogens are rated on two characteristics – invasiveness and toxigenicity. Invasiveness is a measure of the bacterium's ability to grow inside the host, and toxigenicity measures the capacity of the bacterium to produce toxins (chemical substances that cause damage to the host). The combination of these two characteristics gives the final rating of the bacteria's virulence (ability to cause disease). A

species does not necessarily need to have both high invasiveness and high toxigenicity to be rated highly virulent. One or the other can be high enough to cause the bacterium to be very virulent. For example, the bacterium Streptococcus pneumoniae (causes pnuemonia) does not produce a toxin, but it is so highly invasive that it causes the lungs to fill up with fluid from the immune response. In contrast, the bacteria Clostridium tetani (causes tetanus) is not very invasive, but it produces a potent toxin that causes damage at a very small concentration. How does the body fight off a bacterial infection? Again, the body mounts an immune response to the foreign invader, producing antibodies for immediate help and future protection. Since this process takes about a week, antibiotics are usually employed in the meantime. Antibiotic drugs are usually only successful in treating bacterial infections, not viral, or fungal infections. Professionals are becoming concerned that the overuse of antibiotics when they are not needed may lead to the mutation of normal bacteria into antibiotic-resistant bacteria. Bacteria are very resilient and have already developed resistance to many antibiotics. Another concern is that the helpful bacteria that live in the digestive tract may also fall prey to the antibiotics. These bacteria, known as "natural flora," produce vitamins that the host organism uses and needs, as well as help in the digestion of food.

What is a fungus? Fungi (plural for fungus) are different from both viruses and bacteria in many ways. They are larger, plant-like organisms that lack chlorophyll (the substance that makes plants green and converts sunlight into energy). Since fungi do not have chlorophyll to make food, they have to absorb food from whatever they are growing on. Fungi can be very helpful – brewing beer, making bread rise, decomposing trash – but they can also be harmful if they steal nutrients from another living organism. When most people think of fungi they picture the mushrooms that we eat. True, mushrooms are important fungi, but there are other forms such as molds and yeasts. Structure: The main identifying characteristic of fungi is the makeup of their cell walls. Many contain a nitrogenous substance known as "chitin," which is not found in the cell walls of plants, but can be found in the outer shells of some crabs and mollusks. Most fungi are multicellular (made up of many cells), with the exception of the yeasts. The cells make up a network of branching tubes known as "hyphae," and a mass of hyphae is called a "mycelium." The insides of the cells look a little different than bacterial cells. First of all, the genetic material is gathered together and enclosed by a membrane in what is called the "nucleus." Also, there are other structures called "organelles" in the cell that help the cell to function, such as mitochondria (converts energy), endoplasmic reticulum (ER) (makes complex proteins), and other organelles. The Golgi apparatus forms many types of proteins and enzymes. Lysosomes contain enzymes and help digest nutrients. Centrioles are necessary for proper division of the cell. Both bacteria and fungi have ribosomes, but those of the bacteria are smaller in size and also reproduce differently. Reproduction: Fungi can reproduce in multiple ways depending upon the type of fungus and the environmental conditions:

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Budding Fragmentation Production of spores asexually Production of spores sexually

Budding occurs in yeasts, which are only made up of one cell. Budding is somewhat similar to binary fission in bacteria, in that the single cell divides into two separate cells. Fragmentation is a mode of reproduction used by those fungi that form hyphae. During fragmentation, some of the hyphae break off and simply start growing as new individuals. Spores are tiny single cells that are produced by fungi that have hyphae. They can be produced asexually by a process in which the tips of the hyphae form specially encased cells – the spores. Some fungi also produce spores sexually. Two types of special cells called "gametes" are produced. One of each type unite to produce a new individual spore. Spores are tiny single cells that are usually

very resistant to environmental changes. They can remain dormant for long periods of time until the conditions are right for them to develop into mature individuals. Hosts and resistance: Fungi are heterotrophs, meaning that they secrete digestive enzymes and absorb the resulting soluble nutrients from whatever they are growing on. For this reason they are great decomposers in the ecosystem, but they can also cause problems when they begin to absorb nutrients from a living organism. They most commonly are breathed in or have contact with the skin. If conditions are right and they start to reproduce, disease can result. Some antifungal agents are available to treat these infections, but it has been much more difficult for scientists to create successful antifungal drugs than antibacterial drugs because the cells of fungi are much closer in structure to the cells of animals than are bacteria. In creating drugs, it is hard to find an agent that will kill the fungal cells and leave the animal cells unharmed. The most successful drugs that have been created prevent the formation of chitin, and therefore prevent the fungus from creating new cell walls and spreading. The cell wall is the only structure that is not shared by the animal and fungal cells. Other drugs bind to specific fungal proteins and prevent growth. Unfortunately, many of the drugs available are only fungistatic, meaning they can only prevent further growth rather than fungicidal, meaning to kill the fungus. Many of the drugs used for serious fungal infections have potentially toxic side effects.

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Capsule - Some species of bacteria have a third protective covering, a capsule made up of polysaccharides (complex carbohydrates). Capsules play a number of roles, but the most important are to keep the bacterium from drying out and to protect it from phagocytosis (engulfing) by larger microorganisms. The capsule is a major virulence factor in the major disease-causing bacteria, such as Escherichia coli and Streptococcus pneumoniae. Nonencapsulated mutants of these organisms are avirulent, i.e. they don't cause disease. Cell Envelope - The cell envelope is made up of two to three layers: the interior cytoplasmic membrane, the cell wall, and -- in some species of bacteria -- an outer capsule. Cell Wall - Each bacterium is enclosed by a rigid cell wall composed of peptidoglycan, a protein-sugar (polysaccharide) molecule. The wall gives the cell its shape and surrounds the cytoplasmic membrane, protecting it from the environment. It also helps to anchor appendages like the pili and flagella, which originate in the cytoplasm membrane and protrude through the wall to the outside. The strength of the wall is responsible for keeping the cell from bursting when there are large differences in osmotic pressure between the cytoplasm and the environment. Cell wall composition varies widely amongst bacteria and is one of the most important factors in bacterial species analysis and differentiation. For example, a relatively thick, meshlike structure that makes it possible to distinguish two basic types of bacteria. A technique devised by Danish physician Hans Christian Gram in 1884, uses a staining and washing technique to differentiate between the two forms. When exposed to a gram stain, gram-positive bacteria retain the purple color of the stain because the structure of their cell walls traps the dye. In gramnegative bacteria, the cell wall is thin and releases the dye readily when washed with an alcohol or acetone solution.

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Cytoplasm - The cytoplasm, or protoplasm, of bacterial cells is where the functions for cell growth, metabolism, and replication are carried out. It is a gellike matrix composed of water, enzymes, nutrients, wastes, and gases and contains cell structures such as ribosomes, a chromosome, and plasmids. The

cell envelope encases the cytoplasm and all its components. Unlike the eukaryotic (true) cells, bacteria do not have a membrane enclosed nucleus. The chromosome, a single, continuous strand of DNA, is localized, but not contained, in a region of the cell called the nucleoid. All the other cellular components are scattered throughout the cytoplasm. One of those components, plasmids, are small, extrachromosomal genetic structures carried by many strains of bacteria. Like the chromosome, plasmids are made of a circular piece of DNA. Unlike the chromosome, they are not involved in reproduction. Only the chromosome has the genetic instructions for initiating and carrying out cell division, or binary fission, the primary means of reproduction in bacteria. Plasmids replicate independently of the chromosome and, while not essential for survival, appear to give bacteria a selective advantage. Plasmids are passed on to other bacteria through two means. For most plasmid types, copies in the cytoplasm are passed on to daughter cells during binary fission. Other types of plasmids, however, form a tubelike structure at the surface called a pilus that passes copies of the plasmid to other bacteria during conjugation, a process by which bacteria exchange genetic information. Plasmids have been shown to be instrumental in the transmission of special properties, such as antibiotic drug resistance, resistance to heavy metals, and virulence factors necessary for infection of animal or plant hosts. The ability to insert specific genes into plasmids have made them extremely useful tools in the fields of molecular biology and genetics, specifically in the area of genetic engineering. •

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Cytoplasmic Membrane - A layer of phospholipids and proteins, called the cytoplasmic membrane, encloses the interior of the bacterium, regulating the flow of materials in and out of the cell. This is a structural trait bacteria share with all other living cells; a barrier that allows them to selectively interact with their environment. Membranes are highly organized and asymmetric having two sides, each side with a different surface and different functions. Membranes are also dynamic, constantly adapting to different conditions. Flagella - Flagella (singular, flagellum) are hairlike structures that provide a means of locomotion for those bacteria that have them. They can be found at either or both ends of a bacterium or all over its surface. The flagella beat in a propeller-like motion to help the bacterium move toward nutrients; away from toxic chemicals; or, in the case of the photosynthetic cyanobacteria; toward the light. Nucleoid - The nucleoid is a region of cytoplasm where the chromosomal DNA is located. It is not a membrane bound nucleus, but simply an area of the cytoplasm where the strands of DNA are found. Most bacteria have a single, circular chromosome that is responsible for replication, although a few species do have two or more. Smaller circular auxiliary DNA strands, called plasmids, are also found in the cytoplasm. Pili - Many species of bacteria have pili (singular, pilus), small hairlike projections emerging from the outside cell surface. These outgrowths assist the bacteria in attaching to other cells and surfaces, such as teeth, intestines, and rocks. Without pili, many disease-causing bacteria lose their ability to infect because they're unable to attach to host tissue. Specialized pili are used for conjugation, during which two bacteria exchange fragments of plasmid DNA.

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Ribosomes - Ribosomes are microscopic "factories" found in all cells, including bacteria. They translate the genetic code from the molecular language of nucleic acid to that of amino acids—the building blocks of proteins. Proteins are the molecules that perform all the functions of cells and living organisms. Bacterial ribosomes are similar to those of eukaryotes, but are smaller and have a slightly different composition and molecular structure. Bacterial ribosomes are never bound to other organelles as they sometimes are (bound to the endoplasmic reticulum) in eukaryotes, but are free-standing structures distributed throughout the cytoplasm. There are sufficient differences between bacterial ribosomes and eukaryotic ribosomes that some antibiotics will inhibit the functioning of bacterial ribosomes, but not a eukaryote's, thus killing bacteria but not the eukaryotic organisms they are infecting.

Instrument

Uses

Incubator

used for bacterial or fungal cultures

Tyndallizer

a process of sterilization from spore-bearing bacteria

Vaccine bath

used to heat vaccine containing medium gently (to

around 45-55 degrees Celsius) during vaccine production

Inoculation loop:

used to inoculate test samples into culture media for bacterial or fungal cultures, antibiograms, etc.

•Nichrome wire loop

used to inoculate test samples into culture media for bacterial or fungal cultures, antibiograms, etc.; reheated by flaming to red hot before use

•Platinum wire loop

used to inoculate test samples into culture media for bacterial or fungal cultures, antibiograms, etc.; reheated by flaming to red hot before use

•Sterile loops

used to inoculate test samples into culture media for bacterial or fungal cultures, antibiograms, etc.; not heated before use—these are disposable presteriliised

Petri dish/agar plate

to act as a supporting container to hold the culture medium in

production of anaerobic conditions for organisms that die McIntosh and Filde's anaerobic in the presence of even little oxygen (anaerobiosis), eg. jar tetanus bacteria

Gas-pak

releases gases to remove oxygen from a closed container, usually for anaerobiosis

Vacuum pump

to draw out the air from any closed chamber before pumping back CO2, O2 or N2, usually for anaerobiosis

Durham's tube

used to detect gas production in sugar fermentation media; the tube is placed in an inverted fashion so that gases produces get trapped in it and do not float away to the surface

Bijou bottle

a cylindrical small glass bottle with a screw cap used as a culture medium holder

Blood collection bottle

to collect blood by venipuncture

Castaneda's medium / Castaneda's bottle

used for simultaneous solid and liquid cultures in one bottle

Universal container

a cylindrical small glass bottle with a screw cap used as a culture medium holder

Flat medical bottle or McCartney's bottle

for simultaneous solid and liquid cultures.

Tuberculin syringe

as a normal syringe or to perform Mantoux test

Desiccator

to dry things

Pre-sterilized disposable container

specimen collection

Pre-sterilized disposable specimen collection syringe / auto-destruct syringes Pre-sterilized disposable swabs specimen collection / NIH swab / postnasal swab VDRL rotator

for VDRL test

Serological test slides like those for ASO, VDRL, rheumatoid vide links factor

Lovibond comparator

a type of a colorimeter

Microtitre plates

for ELISA

Haemagglutination plate

for viral culture detection

Latex agglutination tiles

for serological analysis

Cragie tube

see link

Tissue culture bottles

to grow or keep alive cells or tissue from a living organism, eg. stem cells

Candle jar

historically used for anaerobiosis; a lit candle was placed in as air-tight jar such that when it went out it would be because it used up all the available oxygen

Optical microscopes Main article: Optical microscope Optical microscopes, through their use of visible wavelengths of light, are the simplest and hence most widely used type of microscope. Optical microscopes typically use refractive glass and occasionally of plastic or quartz, to focus light into the eye or another light detector. Mirror-based optical microscopes operate in the same manner. Typical magnification of a light microscope, assuming visible range light, is up to 1500x with a theoretical resolution limit of around 0.2 micrometres or 200 nanometers. Specialized techniques (e.g., scanning confocal microscopy, Vertico SMI) may exceed this magnification but the resolution is diffraction limited. Using shorter wavelengths of light, such as the ultraviolet, is one way to improve the spatial resolution of the microscope as are techniques such as Near-field scanning optical microscope.

A stereo microscope is often used for lower-power magnification on large subjects. Various wavelengths of light, including those beyond the visible range, are sometimes used for special purposes. Ultraviolet light is used to enable the resolution of smaller features as well as to image samples that are transparent to the eye. Near infrared light is used to image circuitry embedded in bonded silicon devices as silicon is transparent in this region. Many wavelengths of light, ranging from the ultraviolet to the visible are used to excite fluorescence emission from objects for viewing by eye or with sensitive cameras. Phase contrast microscopy is an optical microscopy illumination technique in which small phase shifts in the light passing through a transparent specimen are converted into amplitude or contrast changes in the image. A phase contrast microscope does not require staining to view the slide. This microscope made it possible to study the cell cycle. The Digital microscope appeared a few years ago, using optics and a charge-coupled device (CCD) camera to output a digital image to a monitor.

[edit] Electron microscopes Main article: Electron microscope Three major variants of electron microscopes exist: •

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Scanning electron microscope (SEM): looks at the surface of bulk objects by scanning the surface with a fine electron beam and measuring reflection. May also be used for spectroscopy. See also environmental scanning electron microscope Transmission electron microscope (TEM): passes electrons completely through the sample, analogous to basic optical microscopy. This requires careful sample preparation, since electrons are scattered so strongly by most materials.This is a scientific device that allows people to see objects that could normally not be seen by the naked or unaided eye. Scanning Tunneling Microscope (STM): is a powerful technique for viewing surfaces at the atomic level.

The SEM and STM can also be considered examples of scanning probe microscopy.

Scanning acoustic microscopes use sound waves to measure variations in acoustic impedance. Similar to Sonar in principle, they are used for such jobs as detecting defects in the subsurfaces of materials including those found in integrated circuits.