lab_03_aseptic_technique

lab_03_aseptic_technique - Miramar College Biology 205...

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Lab Exercise 3: Aseptic Technique Page 1 of 3 Miramar College Biology 205 Microbiology Lab Exercise 3: Aseptic Technique Background Aseptic technique is one of the most fundamental techniques in all of microbiology. The ability to keep your cultures contamination-free is as important as keeping you from being contaminated- for different reasons, of course. When you are attempting to identify an unknown organism, whether from a patient or an environmental sample, or you’re trying to characterize an organism that you’ve already identified- having a pure culture of that organism is a must. The only way to achieve these pure cultures is by practicing aseptic technique, the transfer of microbial cultures from one medium to another without introducing contamination or damaging the organism of interest. Sterilization is typically achieved by one of two methods: dry or moist heat . Typically in a microbiology lab dry heat sterilization is achieved by flaming your instrument in the flame of a Bunsen burner. Moist heat is most often achieved through the use of an autoclave . In the microbiology lab, you will typically use media that have been autoclaved to sterilization and you will maintain this sterility by dry heat sterilizing every instrument that you use which comes into contact with the media. In this way, you will avoid any contamination and any resultant growth will be only the organism which you have deliberately inoculated into the medium. Depending on your needs, bacteria can be cultured onto different types of media. For routine growth, oftentimes inoculation into a broth culture is best. For isolating specific bacteria from a mix, growth on a Petri dish is optimal. For long-term storage and maintenance of a culture, growth on an Agar slant is best and for anaerobic growth, inoculation into an Agar deep is optimal. Introduction Media Media comes in both liquid ( broth ) as well as solid ( agar ) form. In order for media to be a solid, a solidifying agent (most commonly agar) is added to the broth prior to sterilization. Solid media is prepared into Petri plates or in tubes as slants or deeps (Figure 1).. Additionally, a pour can be made by pouring molten agar into a Petri dish. Figure 1: Different types of media inoculations. Sterilization and the Autoclave When microbiological media has been made, it still has to be sterilized because of microbial contamination. Sterilization, by definition, kills all organisms in the media and will guarantee that the medium will stay sterile unless it is handled incorrectly (to be explained further in the Aseptic Technique portion of this laboratory exercise).
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