My cells were supplemented with 0.5% fetal bovine serum (FBS), 10 ng/mL of epidermal growth factor, 5 mg/mL of insulin, 0.5 mg/mL of hydrocortisone, 0.5 mg/mL of epinephrine, 6.5 mg/mL of triiodothyronine, 10 mg/mL of transferrin, 10 mg/mL of gentamicin, and 50 ng/mL of amphotericin-B.
I have only one vial left in liquid nitrogen, Please suggest me how to save my last vial?
Note: There is no contamination in other cell lines growing in CO2 incubator but only in HRPTEC line.
I think the only thing you can do is clean everything, thaw the last vial and hope for the best that your stock was not contaminated. You should not forget that prolonged treatment of the cells with combinations of antibiotics may have an impact on your results.
You should discard every plastic cell culture material you have used so far and all mediums, trypsin, PBS, etc. Make new medium, take new aliquots of everything you use, new PBS, new trypsin, etc. Disinfect the hood area before work with 75% ethanol, use UV light 15 min before work, use protection coat and wear gloves. Every time before putting something in the hood, disinfect it with ethanol. When everything is set, thaw your last vial and hope the cells in it were not contaminated before freezing!
Good luck.
Thanks a lot. I did exactly similar and my cells are good now. I'm about to freeze them.
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