The spread plate isolation method is a procedure where the researcher spreads a diluted cell mixture on an agar surface, using a sterile bent glass rod, in a way that every cell grows into its own separate colony. The cultures are incubated for a specific period of time. A macroscopically visible growth or cluster of microorganisms on the solid medium appears, each colony represents a pure culture. After colonies are grown, they are counted and the number of bacteria in the original sample is calculated.
Pour plate procedure is extensively used with bacteria and fungi and can also yield isolated colonies. A diluted bacterial specimen is prepared. The researcher introduces a small amount into an empty Petri dish. Melted agar is poured into the Petri dish containing the specimen. Agar and specimen are mixed thoroughly by covering the dish and gently tilting and swirling it. Agar is allowed time to gel while sitting on a flat surface. The covered dishes are inverted and incubated for a specific time period. Colonies are then observed and counted, and information is recorded.
The streak plate method is the classic procedure for isolating individual strains of bacteria. The researcher sterilizes a wire loop in a flame. After cooling, the wire loop is touched to a sterile agar plate. The researcher dips the loop into the sample, picking up thousands of bacteria, then streaks the loop back and forth across the agar surface. After sterilizing the loop again and allowing it to cool, the researcher drags the loop through the previous path, picking up a batch of bacteria and streaks it onto the agar in a new area. This procedure is repeated several times. Closed dishes are incubated for a specific time period, after which the colony growth is observed.
Exposure of the sterile media to the environment demonstrates the aseptic technique. The researcher labels two nutrient agar plates for later comparison and observation and leaves them uncovered. One exposed agar plate is allowed to remain just as it is, exposing it to the environment for a short time period. The second plate is exposed to a source of possible contaminants, such as a sneeze, cough or fingers placed in physical contact with the agar surface. The plates are covered, inverted and incubated at room temperature for a period of time before observation.