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Encyclopedia > Agar plates

An agar plate is a sterile Petri dish that contains agar plus nutrients, and is used to culture bacteria or fungi. Man looking at fungus inside of petri dishes A Petri dish is a shallow glass or plastic cylindrical dish that biologists use to culture microbes. ... Agar is a galactose polymer (or Agarose) obtained from the cell walls of some species of red algae or seaweeds (Sphaerococcus euchema) and species of Gelidium, chiefly from eastern Asia and California. ... Phyla/Divisions Actinobacteria Aquificae Bacteroidetes/Chlorobi Chlamydiae/Verrucomicrobia Chloroflexi Chrysiogenetes Cyanobacteria Deferribacteres Deinococcus-Thermus Dictyoglomi Fibrobacteres/Acidobacteria Firmicutes Fusobacteria Gemmatimonadetes Nitrospirae Omnibacteria Planctomycetes Proteobacteria Spirochaetes Thermodesulfobacteria Thermomicrobia Thermotogae Bacteria (singular, bacterium) are a major group of living organisms. ... Divisions Chytridiomycota Zygomycota Ascomycota Basidiomycota The Fungi (singular: fungus) are a large group of organisms ranked as a kingdom within the Domain Eukaryota. ...

Contents


Preparation of agar plates

Most types of agar are purchased in powder form. They are dissolved in distilled water as per their instructions. It usually necessary to gently boil the mixture to facilitate dissolving: this can be done in a microwave oven, or over a gentle flame. Once dissolved the agar needs to be sterilised, usually by pouring it into a Erlenmeyer flask, then sealing the top with a cotton wool wad, and finally covering the cotton wool with a loose layer of aluminium foil. This is then autoclaved for 15 minutes. The sterile agar is then allowed to cool to 50 °C: this is just above the setting point of agar and pouring at this cooler temperature helps prevent condensation forming on the lid. Erlenmeyer flask An apparatus feeding into an Erlenmeyer flask An Erlenmeyer flask (also called conical flask) is a type of laboratory glassware which consists of an inverted conical base with a cylindrical neck. ... Strictly, Cotton wool is the silky fibers from cotton plants in their raw state. ... Front loading autoclaves are common Cassette style autoclaves cycle very quickly Stovetop autoclaves need to be monitored carefully, but have a very large capacity Sterilization bags often have a “sterilization indicator mark” that typically darkens when sterilization temperatures have been reached. ... Condensation can refer to: The change in phase of a substance to a denser phase, such as gas to a liquid. ...


Before the plates are poured, every care is taken not to contaminate them with stray bacteria: sterile technique must be used.

  1. Hands are thoroughly washed with antimicrobial soap and hot water.
  2. The bench is wiped with ethanol or some other disinfectant
  3. A Bunsen burner is set up with a gentle blue flame. This will be used to sterilise the mouth of the flask, and will also provides a reasonably sterile environment in the vicinity. This will also be used to flame plates that develop bubbles from pouring.
  4. The number of plates to be poured are placed on the bench, with their lids still on.
  5. The aluminium foil is removed and discarded. The cotton wool is removed with the little finger. It is held in the little finger the whole time, not put down.
  6. The mouth of the flask is flamed to kill bacteria on the outside of the rim.
  7. The lid of the plate is lifted just high enough to allow the plate to be poured, and the dish is quickly half filled with agar.
  8. The lid is replaced, the plate swirled gently to ensure even distribution of the molten agar, then left to stand on the bench for at least 20 minutes to solidify.
  9. Once all the plates are poured, the flask mouth is reflamed and the cotton wool reinserted. Any unused agar is still sterile.

To allow traceability, a lot number assigned to the flask is written on the plates poured with it. Antibacterial soap is any cleaning product to which purportedly antibacterial chemicals have been added. ... Ethyl alcohol, also known as ethanol or grain alcohol, is a flammable, colorless chemical compound, one of the alcohols that is most often found in alcoholic beverages. ... Disinfection The destruction of pathogenic and other kinds of microorganisms by physical or chemical means Disinfectants are chemical substances used to kill viruses and microbes (germs), such as bacteria and fungi. ... A bunsen burner with needle valve. ...


Inoculation techniques

Before inoculation, important information is written on the bottom of the plates, close to the rim:

  1. date of inoculation
  2. temperature of incubation
  3. duration of incubation
  4. microorganism inoculated

Streaking

The most common method of inoculating an agar plate is streaking.

Image:Streak plates.png
Streak plates
  1. With this method, a small amount of sample is placed on the side of the agar plate (either with a swab, or as a drop from an inoculating loop if the sample is a liquid).
  2. A sterile loop (flamed until red hot, then cooled by touching the agar away from the inoculated sample) is then used to spread the bacteria out in one direction from the initial site of inoculation. This is done by moving the loop from side to side, passing through the initial site.
  3. The loop is then sterilised (by flaming) again and the first streaks are then spread out themselves.
  4. This is repeated 2-3 times, moving around the agar plate.

What should happen is that single bacterial cells get isolated by the streaking, and when the plate is incubated, the resulting colonies will each have started from just one bacterium.


Christmas tree

This pattern is used for culture of urine. A small loop is dipped in the urine, and a single streak is made down the middle of the agar plate. Then the loop is swayed in and out going through the streak multiple times at right-angles to the first streak. Urine is liquid waste excreted by the kidneys and eventually expelled from the body in a process known as urination. ... This article is about angles in geometry. ...


Stab culture

A needle is flamed then immersed in the culture. It is then stabbed into a small sterile jar of nutrient agar. If the bacteria are anaerobic they will grow, otherwise they do not. An anaerobic organism or anaerobe is any organism that does not require oxygen. ...


Preparing a lawn

A lawn is often used for testing sensitivity to antibiotics, or for work with bacteriophages. What is needed is an even and complete spread of growth all over the agar plate (a "lawn"). Around an antibiotic disc there will be a clear area in which bacterial growth is inhibited; the diameter of this area can be measured to find out whether that bacterial strain is resistant to the antibiotic. An antibiotic is a drug that kills or slows the growth of bacteria. ... A phage (also called bacteriophage) (in Greek phageton = food/consumption) is a small virus that infects only bacteria. ... An antibiotic is a drug that kills or slows the growth of bacteria. ...


One way to prepare a lawn is to use a 0.5 McFarland suspension of bacteria in saline (this means the saline is made just slightly turbid.) A sterile swab is dipped into this suspension, then it is moved from side to side down the whole agar plate so all the area is covered. The plate is rotated 90 ° and the swab moved side to side perpedicularly to the first time. This is done once more with the swab rotated 45 °. In microbiology, McFarland standards are used as a reference to adjust the turbidity of bacterial suspensions so that the number of bacteria will be within a given range. ...


Once a lawn has been prepared, a small disk of sterile filter paper is soaked in antibiotic and placed on the plate. After incubation there will be a ring of zero growth visible around the filter paper if the lawn bacteria are sensitive to that antibiotic. A collection of small disks each soaked in a different antibiotic, and attached to a larger ring, can be purchased commercially. They are known as antibiotic sensitivity discs or by the name "Mastrings" (a trademark of Mast Group Ltd) and can be used to test the sensitivity of an organism to a range of antibiotics all at once.


Incubation of agar plates

Plates are incubated upside down to prevent drops of condensation from collecting on the inoculated surface.


Most plates are incubated at 37° C in a 5% CO2 atmosphere: the temperature and conditions that most of the body's bacteria will grow. Special incubators can maintain these conditions.


Some bacteria must be incubated anaerobically (without any oxygen). These can be placed in containers, along with a substance that removes oxygen, and the tightly sealed container placed in the regular incubator.


Fungi, and some bacteria (e.g. Yersinia sp.), should be incubated slightly cooler. This is usually 30 °C, and room air often is used.


Yoghurt bacteria grow at much higher temperatures (typically ~45 °C). They are therefore particulally safe to use when teaching microbiology, especially to children.


Campylobacter is a difficult bacterial species to grow. It needs special agar plates, plus its own microaerophilic environment. Species see text Campylobacter is a genus of Gram_negative bacteria. ...


Types of agar plates

  • Blood agar - contains blood cells from an animal (e.g. a sheep). Most bacteria will grow on this medium.
  • Chocolate agar - this contains lysed blood cells, and is used for growing fastidious (fussy) respiratory bacteria.
  • Neomycin agar - contains the antibiotic neomycin.
  • Sabouraud agar - used for fungi. It contains gentamicin and has a low pH that will kill most bacteria.
  • Thayer-Martin agar - chocolate agar designed to isolate Neisseria gonorrhoeae.
  • XLD agar - xylose lysine deoxycholate agar. It is used for the culture of stool samples, and contains two indicators. It is formulated to inhibit Gram-positive bacteria, while the growth of Gram-negative bacilli is encouraged. The colonies of lactose fermenters appear yellow.
  • Nutrient agar - safe to use in school science laboratories because it does not selectively grow pathogenic bacteria.

Binomial name Ovis aries Linnaeus, 1758 A sheep is any of several woolly ruminant quadrupeds, but most commonly the Domestic Sheep (Ovis aries), which probably descends from the wild moufflon of south-central and south-west Asia. ... Neomycin is an antibiotic that is found in many topical medications such as creams, ointments and eyedrops. ... Raymond Sabouraud (November 24, 1864 - 1938) was a French physician specialized in dermatology but also a painter and sculptor. ... Gentamicin is a aminoglycoside antibiotic, and can treat many different types of bacterial infections, particularly Gram-negative infection. ... pH is a measure of the activity of hydrogen ions (H+) in a solution and, therefore, its acidity or alkalinity. ... Neisseria gonorrhoeae is a species of Gram-negative (see also Gram Stain) bacteria responsible for the disease gonorrhoea. ... A stool can refer to: A type of chair feces This is a disambiguation page — a navigational aid which lists other pages that might otherwise share the same title. ... Gram-positive bacteria are those that are stained dark blue or violet by gram staining, in contrast to gram-negative bacteria, which are not affected by the stain. ... Bacteria that are Gram-negative are not stained dark blue or violet by Gram staining, in contrast to Gram-positive bacteria. ... Orders Bacillales Lactobacillales The term bacilli (singular bacillus) is used to refer to any rod-shaped bacteria. ... A pathogen (literally birth of pain from the Greek παθογένεια) is a biological agent that can cause disease to its host. ...

Safe disposal of agar plates

Plates, once finished with, must be made safe before throwing away. The usual method is to place inside an autoclave bag and then sterilise by autoclaving at 121 °C, 103 kPa (15 psi) for 15 minutes. Plastic plates will melt (hence the bag). After about 20 minutes the autoclave will have cooled down and the bag can safely be thrown away. If no autoclave is available an ordinary domestic pressure cooker can be used, or, in a hospital or professional lab an incinerator may be used. Front loading autoclaves are common Cassette style autoclaves cycle very quickly Stovetop autoclaves need to be monitored carefully, but have a very large capacity Sterilization bags often have a “sterilization indicator mark” that typically darkens when sterilization temperatures have been reached. ... The pascal (symbol Pa) is the SI unit of pressure. ... Psi has multiple meanings: Psi (Ψ ψ) is a letter of the Greek alphabet. ... Pressure cooking is a method of cooking things at high heat without boiling them. ... This article needs cleanup. ...


Other equipment may be decontaminated by being placed in a suitable disinfectant such as Virkon for 24 hours. Virkon is a brand name of a powerful disinfectant, often used in laboratories for cleaning up spills, soaking equipment or wiping benches. ...


When all manipulations are done, the bench is disinfected once again. The last step should be to wash the hands thoroughly with antimicrobial soap and hot water before leaving the lab.


See also

Laboratory equipment
Agar plate | Aspirator | Bunsen burner | Calorimeter | Colorimeter | Centrifuge | Fume hood | Microscope | Microtiter plate | Plate reader | Spectrophotometer | Thermometer
Laboratory glassware
Beaker | Boiling tube | Büchner funnel | Burette | Conical measure | Cuvette | Laboratory flasks (Erlenmeyer flask | Florence flask | Volumetric flask | Buchner flask) | Gas syringe | Graduated cylinder | Pipette | Petri dish | Separating funnel | Soxhlet extractor | Test tube | Thistle tube | Watch glass

  Results from FactBites:
 
Agar plate - Wikipedia, the free encyclopedia (598 words)
An agar plate streaked with microorganisms isolated from a deep-water sponge.
A type of blood agar plate in which the blood cells have been lysed by heating the cells to 56 °C; used for growing fastidious (fussy) respiratory bacteria.
HE agar is designed to isolate and recover fecal bacteria belonging to the Enterobacteriaceae family.
Agar - Wikipedia, the free encyclopedia (784 words)
Chemically, agar is a polymer made up of subunits of the sugar galactose; it is a component of the algae's cell walls.
Research grade agar is used extensively in plant biology as it is supplemented with a nutrient and vitamin mixture that allows for seedling germination in petri dishes under sterile conditions (given that the seeds are sterilized as well).
Nutrient agar is used throughout the world as a medium for the growth of bacteria and fungi, but not viruses (however, viruses are often grown in bacteria that are growing on agar).
  More results at FactBites »

 
 

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