I have been doing my scaffold research which makes use of C2C12 to test biocompatibility. Seeding density was at 1x10^5 and 5X10^5 cells/5mm disk (0.2 cm^2) with proliferation media changed everyday. The earliest recorded contraction was 3 days after seeding. Passaging of cells prior to seeding in scaffolds was done at 80-90% confluency via trypsinization. I have 2 hypotheses: 1) early differentiation due to high seeding density & 2) effect of scaffold (release of bioactive components that could promote improved proliferation leading to early differentiation).
I am not sure if it's differentiation even if it's confluence. You can check the differentiation markers. Even in the serum free media it take 7days to differentiate to myotube. As it's very easy to check, I suggest you can just check the status of the genes with serum free media after 7days and 3days with normal medium cultured on scaffold to confirm your hypothesis. As you can also have potential finding related to the scaffold effect on the C2C12 cells. I think it's worth a try. Good luck and best wishes
Zannatul Ferdous contractions were also recorded starting from the 3rd day of proliferation.
multiple contracting myotubes were recorded on the 7th day of proliferation via electrostimulation. I also did western blot which gave positive detection of MF20 and MYO5A
That seems great. How about conducting PCR with MYOD, Myogenin, MYO5, MYF4? Better to check the status of differentiation. As you may already know there are several states of differentiation in case of Myoblast cells.
I should add that to my future experiments. in the mean time, i am limited to protein level only (No primers yet). In my opinion, myotube contraction is a sign of terminal differentiation of myoblasts.
- Badges
- Science topic