Greenville, South Carolina
AUTOCLAVE: LEARNING HOW TO KILL (STERILIZATION)
An autoclave reaches 121 C and 15psi. Therefore it kills nearly any bacteria that could cause contamination. Without the sterilization, NO microbiology experiment would work. To achieve sterilization we have to learn how to use an autoclave.
The use of an autoclave serves three purposes ? to sterilize media, to sterilize glassware and to sterilize biohazardous wastes.
The following instructions are for use of the two autoclaves in Peale Science Center at Hope College.
USING THE AUTOCLAVE TO STERILIZE MEDIA
[images of flasks with liquid media]
The amount of medium you need to prepare depends on what you will be using it for. As we mentioned before, there are two types of media, solid and liquid. All media will be autoclaved and therefore as it comes out of the HOT autoclave all media will be in the liquid state. If the media contains agar, it will harden while it cools. This type of media is used to pour petri plates. The amount of media you makes depends on how many plates you need. Each Petri dish holds about 20-30ml of medium. (More on pouring plates in section 7).
If you are making media in which to grow bacteria you will probably need to aliquot portions of it into tubes. Growing bacteria in liquid media is discussed in depth in section seven, but it is important that you realize you will need different volumes of media (i.e. 10ml tubes, 300ml flasks, 500ml flasks, etc.). Before autoclaving, pour the correct amount of media into the tubes or flasks, which will be utilized in the experiment.
Use the autoclave in the microbiology prep room, 154b.
[images of an autoclave]
USING THE AUTOCLAVE TO STERILIZE GLASSWARE OR OTHER LAB UTENSILS
Autoclaving glassware, pipettes, sticks and other general lab supplies will eliminate the amount of contamination. Use the autoclave in the microbiology prep room, 154b.
USING THE AUTOCLAVE TO STERILIZE TRASH
Biohazard waste treatment is essential in microbiology. Any used plate, kimwipe, or even cotton swab, which has been in contact with living organisms, has the potential to multiply and spread bacteria. Therefore, they must be autoclaved before it can be thrown away in regular trash can.
Use 'kill' autoclave in the greenhouse prep room for trash sterilization.
STERILIZATION EXERCISES: (read through all instructions before you start)
The medium that was autoclaved should be clear, whereas the medium, which was not autoclaved, should be cloudy. The cloudiness is due to bacterial contaminants. Remember, the media you make will satisfy the desires of many different bacteria. Since you will only want the specific bacteria you are working with to grow in the media you make, be sure if is VOID of any other bacteria before using it.
Now that you learned how to sterilize your media you need to learn how to KEEP the unwanted bacteria away from the media enter the aseptic technique.