Hello,
I'm now working on C2C12 myoblast cell line.
Growing the cells was great, but there are some problems during differentiation process.
The cells are not differentiating but frequently dying.
I got C2C12 stock from ATCC.
After I thawed, I grew the cells about a week changing the flast to bigger size, not throwing away any cells.
Then, I made 32 stock for later experiments and one T75 culture (1M cells).
On the day that the confluency reached about 80%, I switched growth media to differentiation media.
However, there is no sign of differentiation but cell death.
I did same process on 100% confluency cells but it seems to be same.
I used growth media (DMEM with high glucose + 10%FBS) and differentiation media (DMEM with high glucose + 2%HS)
Attached file is microscope images (On the day of changing media to Day 6)
As you can see, there is no myotube formation.
What should I change to get better outcome?
Hi,
Was the culture media turbid? From your images, it looks like you have quite a bit of contamination. I first recommend ensuring aseptic technique and consider supplementing your growth and differentiation media with antibiotic (penicillin-streptomycin).
Good luck!
Joshua R Huot Adam P Lightfoot
Thank you for anwsering.
During culture and differentiation process, there was no sign for contamination
(I think the image is messy due to old CCD camera)
I'm now planning to test using various serum cocentrationtion.
I'll try to add antibiotics at that time
Again, Thank you!
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