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Sterile Technique

    General Guidelines
  1. Maintain a clean work area
  2. Use a fresh pipet tip for every transfer (tips should be DNase/RNase free)
  3. Wear gloves to prevent contamination (of yourself as well as your experiment)
  4. Sterilize solids and liquids by autoclaving 20 minutes at 121C at 15 psi

Microcentrifuge/conical centrifuge tubes can easily be contaminated by contact with non-sterile surfaces (e.g., your fingers) or by air borne particles. Be careful when transferring solutions from one tube to another. Also, keep the lids closed when you're not working with the samples.

Media can be also contaminated by contact with non-sterile surfaces or by air borne organisms. Remove lids and coverings carefully avoiding contact with any part of the cover that may contact the media; minimize the amount of time the container is exposed to air. Lids and coverings should be held with media side down at all times. Air borne contaminants are usually falling downward. Replace the coverings carefully so that the rim of the container makes contact only with sterile surface of the inside of the cap.
The use of a flame helps maintain aseptic materials. Working near a flame can decrease air borne contamination. The flame is also used to singe surfaces to maintain sterility. The mouth of the tube or flasks is passed through the flame before and after pouring. The cap or cover is also passed through the flame prior to replacing on the container.
Caution: The flame is used to singe the surfaces only. Do not hold the items in the flame to make them hot. Glass flasks, even Pyrex, can break from the heat or when the cooler media hits the hot surface.

For additional suggested procedures, see Appendix: Sterile Technique, etc.


Copyright, Acknowledgements, and Intended Use
Created by B. Beason (bbeason@rice.edu), Rice University, 6 February 2008
Updated 7 February 2008