I have to study cell cycle analysis using my miRNA inhibitor. Is it necesaary to synchronize cells before doing FACS? what is the protcol for it?
Not all cell-types can be synchronized using serum starvation because certain cell-types undergo cell death. Therefore, if you are considering to serum starve normal fibroblasts or epithelial cells, try starving for 24 or 48 h. This starvation period is likely to result in accumulation of >90% cells in the G0/G1 phase. The next question would be whether the inhibitor would delay entry into the S-phase.
Thanks Dr. Choubey. I am using hepatocarcinoma huh7 cells. I am using miRNA-106b inhibitor which targets Rb in my study. i read papers where they have used serum starvation for cell synchronization in huh7 cells.
I believe that 48 h serum starvation may work for your cell line. If not try 0.2% serum.
Thanks Dr. Choubey. 24 hours of serum starvation are killing my cells with 1% serum. when i am preparing these samples for FACS, cells are getting clumped.
We currently performed a reduction of serum 72h at 5% and then 72 h at 0% cells at high confluency ;
For studying cells in G0 phase, is it important to do serum starvation before treatments?
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