I made thawing of a cryopreseved cell line, after transferring into t-75 flask and examination under microscope. it showed a contaminated cell culture. The contamination apparently had occurred before cryopresevation. Is there any method to overcome these contaminants.?? For example, increase the concentration of antibiotics or using broad spectrum agents as ciprofloxacin?....Thanks in advance
Better you discard the flasks and start with a new culture.... Otherwise it may contaminate other cell lines too...
First of all: how much yeast and bacteria does it have? Are your cells adherent?
If your cells are adherent and you do want to try to save them, you can try the following protocol:
1) Remove the supernatant and place 4mL HEPES anti/anti (1% solution). Put the t-75 flask in the cell incubator for 15 minutes.
2) Remove the supernatant and repeat the previous step twice more (total - 3x 15min "washes").
3) Feed your cells with medium 1% anti/anti and watch them for the next few days.
Rafael Factor Caran : In fact the cells are partially adherent, I understand that 1% anti/anti is sufficient. Thanks so much.
I agree with Mavis, you should disccard the flask. You could try repeatedly washing them and place antibiotics, but my experience has been that it is very difficult, if not impossible to decontaminate a culture that has become infected.
The only option is to start the culture again. You will waste a lot of time trying to remove contamination and never be sure. Time should be spent on assessing how this happened. Always avoid chances of cross contamination.
Use antibiotics like Panstrap (1x Solution), Amphotericin, Gentamycin and Tylosin as anti-mycoplasma.
Follow simple things to avoid contamination as Prevention is better than cure and this problem has no cure mostly.
All the best.
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