My HGF cell line was working fine when suddenly every cryovial i thawed started to show a pattern of cell death, i.e. after one subculture. Contamination comes out of nowhere and i am losing valuable cell stock. The incubator has been cleaned , all plastic ware changed and reagents changed as well. The aseptic technique is spot on and frankly was working fine before. I am at a loss to understand why this is happening. The media i use is alpha-mem, P-strep 1% and FBS 10%. Filtered FBS stock after throwing away previous and got new P-strep and alpha-mem as well. Any ideas or suggestions because this is hindering my entire study.
Maybe your cells themselves had some degree of contamination when they were frozen and now cells AND the contaminants are being revived on thawing? Grow some other cell line under the same conditions with the same stocks of materials. If they grow well without contamination, then these cells are contaminated.
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