How can I know that HepG2 cells are contaminated? I find their shape abnormal in the TC flask. There are a lot of intracellular small bubbles. Any suggestions?
Promega provides cell line authentication service: http://www.promega.com/products/str-analysis/cell-line-authentication/cell-line-authentication-service/
If contamination is there in the Hep2 cultured cells and any other cultured cells, the tightly adhere cells become loose and simple shaking of the plate might be able to float the cells apart from abnormal shapes. Also, black spots could be visible inside the cells.
That certainly looks like mycoplasma. Unfortunately, the test I liked most - the mycotect kit - has been discontinued. You could use a PCR detection instead or build up a mycotect-like kit yourself with 6-methylpurine deoxyriboside (6-MPDR) and 3T6 cells as indicator cells, if you find the time. However, the easiest way is to simply discard your cells and thaw another sample or get one from another lab. Don't waste your time on a contaminant!
To test for mycoplasm as suggested by Otero and to use as low passage numbers as possible could help to solve the problem. However, I would not use DMEM but instead Eagle's Minimum Essential Medium or RPMI supplemented with 10% FBS, penicilin
and streptomycin (50U/ml) and L-glutamin 1%.
We experienced some time ago that e.g. cultivating the in suspension growing HL60 cell line in DMEM instead of RPMI changes their size and granulyrity greatly and the tend to bind artificially to Annexin V-FITC. Thus media matters a lot.
I am also using HepG2. Cells are not contaminated untill you see any black things.
Intracellular bubbles are when they start getting unhappy basically they are ready to split. Just keep an eye that when they are arround 70 % confluent. Just spilt them and keep doing this untill 3-4 passages. My cells have been seem far better after doing this. If still not statisfied, you can do some staining and see if you have any Mycoplasma as reffered above.
Good Luck
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