Helpful Hints For Better Aseptic Technique

By Bionique Testing Laboratories

Developing successful aseptic techniques requires specific training and a clear understanding of the nature, types, and potential sources of contamination. Freshney (2005) and Ryan (2008a) provide good information on these critical areas. The following recommendations are designed to help improve your aseptic techniques and to preserve the integrity of your cell cultures.

General principles of handling cell cultures: (Freshney; 2006)

• First and foremost, all supplies and reagents that come into contact with the cultures must be sterile (Phelan, 2007).
• Wash hands before and after handling any cell culture. Hand washing stations should be readily accessible within the laboratory.
• Handle only one cell line at a time. There are intrinsic risks of misidentification or cross contamination between cell cultures when more than one cell line is in use within the laboratory (Freshney, 2006).
• Handle continuous cell lines after the handling of short-term, finite cell cultures.
• Quarantine and handle under strict precautions all incoming cell lines until testing concludes the absence of mycoplasma. Alternatively, purchase cell lines from repositories which certify that materials are mycoplasma-free prior to distribution.
• Avoid continuous long-term use of antibiotics within cell cultures. The overuse of antibiotics as prophylaxis may lead to cytotoxicity and pose an increased risk of covert mycoplasma contamination within the cell lines. For further information on this important topic, please refer to FAQ: Why does overuse of antibiotics result in higher mycoplasma contamination rates? (2009; Available within the Mycoplasma Resource Center at www.bionique.com)
• Cultures should be inspected daily for signs of contamination. In addition, testing at regular intervals for mycoplasma should be conducted to ensure the purity and integrity of the culture.
• Promptly discard any contaminated cultures. Retention of these cultures poses a serious threat of cross contamination to other cultures in the laboratory. If clean-up of the contaminated culture is attempted, then any work with this culture should be reserved to the very end of the day to minimize transfer of the contamination.