Sterilization and Aseptic Techniques

• Sterilization - process of eliminating any form of life from any material; done prior to any culture work in the lab
• Steam sterilization - surest and preferred technique; moist heat does the sterilizing not the pressure; temp 121°C and a pressure of 15 PSI for 15 minutes; living organisms die in 10-12 minutes
• Fractional sterilization - for bacteriological media that can withstand a temp of 100°C; 3 successive days for 20 to 30 minutes with incubation period in between
• Boiling - least expensivee and readily available method but not recommended in the lab. Vegetative cells killed within 5 to 10 minutes but spores and certain viruses can survive for many hours of this treatment
• Dry heat by direct flaming or incineration - simplest with the use of hot flame; 100% effective especially in the sterilization of loops and needles for inoculation purpose
• Hot air sterilization - oven-like temp of 160°C to 170°C for 2 hours; has less capacity to penetrate than moist heat; requires longer exposure time to kill all forms of life
• Filtration - sterilize fluid or liquid components that are thermolabile
• To ensure that sterilization had been properly done, it is necessary to test the media, for instance the sterility before inoculation with bacteria or any material.
• Plated media are incubated overnight at 34 to 37°C
• Only those media that do not show any growth of microorganism are considered sterile and can be used.
• Aseptic techniques - procedures that prevent contamination (acquiring undesirable microorganisms)
• Aseptic techniques include:

  Heating the wire loop until red hot

  Flaming the mouth of test tubes/flasks

  Disinfecting your working area with an antiseptic

  Working in a disinfected hood that prevents the flow of contaminated air

  Keeping one's mouth shut while inoculating