i have observed contamination on some of my cell lines, bacterial stain indicated its a coccus type of bacteria, i still need the cells so is there any way to get rid of them??
1.centrifuge cells at 1000 rpm then collect them in clean tube. this will reduce the number of bacteria because most bacteria will be in the supernatant.
2. add tissue culture medium with 2x pencillein/streptomycin + 2x gentamiycin to the cells in the clean tube and centrifuge again
3. culture your cells in new flask with media containing 2x pencillein/streptomycin + 2x gentamiycin.
4. if your cells are adherent, keep changing media daily with gradual increase in antibiotics concentration until the contamination disappear.
are you using human or animal cells?
thank you so much for giving me useful tips Dr. Wamidh...... am using human cancer cell line
Lalrinzuali - If there's any way, I'd try to find clean cells and start over.
That said, I'd amplify what Wamidh suggested above. If you're seeing cocci, then you probably have typical contaminating organisms from human skin (e.g., Staphylococcus epidermidis, etc.). These will typically be very sensitive to pen/strep/gent. If you're finding that it's not working, I'd plate the material on a general use bacterial agar like Mueller Hinton. Subculture every colony morphology that you find, and submit those for culture and sensitivity to a clinical laboratory. That information can provide you detailed information about antibiotic sensitivity. Note that this approach will probably cost a fair amount (at my old University, it would be several hundred dollars to sort this out). If your cell line is really irreplaceable, then trying to understand the cause of the contamination may be worth it.
Best, John
i prepared new media as Dr. Wamidh suggested and centrifuged the cells as well, now i don't see any contamination,however, i have started another culture with new cells after cleaning everything as Dr.Younger suggested as well... thanks to both of you for giving your time to answer my question
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