I am trying to transiently induce EMT in HepG2 cells using TGFb/Activin recombinant proteins. What should be the ideal confluency before starting the treatment? And is serum starvation required prior to the treatment and withdrawn altogether during the EMT induction?
Hi Ain,
While studying EMT induction in HepG2 cells caused by TGF-β1 (without activin), we proceeded as follows. HepG2 were cultured in RPMI containing 10% fetal calf serum with Penicillin/Streptomycin. Medium was renewed every second day. For the experiment, cells were passaged and plated in 6-well dishes AT A DENSITY OF 4 × 105 cells. One day before the experiment, cells were washed with PBS (1×), medium changed to HepatoZYME-SFM and cultured for further 24 hrs. One hour before the experiment, the medium was exchanged and HepG2 cells stimulated with 1 ng/mL recombinant human TGF-β1 for time intervals: 0 min, 20 min, 1 h, 2 h, 4 h, 24 h. The cells were harvested using Qiazol for cell lysis. Serum starvation was NOT required.
Best wishes,
Ilya
- Badges
- Science topic
- Similar topics
- Biological Science
- Molecular Biology
- Biomolecules
- Nucleic Acids
- RNA