Hepg2 transfection - (Jul/20/2007 )
What would be an optimal reagent and protocol to transfect Hepg2 cells? That would be my first cell transfection in this lab. Previously, they were using nucleofector T28 protocol, would there be another good choice for?
Lipofectamine 2000 should a good choice. ~20-30% transfected.
I've read that its cytotoxic to cells. Have you transfected Hepg2 specifically with this system? What advice can you give me for using this system with HepG2 to minimize cell death and increase %?
All transfection reagents are more or less toxic. Hep G2 seems to be more resistant than other cell lines that I have worked with. I dont recall any serious problem with Lipo2000 in these cells. I used 0.2 to 0.3 ug DNA per well for cells in 96 well plate. 1 : 3 to 1 : 5 DNA (ug) to lipo2000 ratios. It will best for you to do a titration on the ratios and cell density in 96 well formate.
Replace medium containing transfection mixture after 5 hrs. Add a low % FBS, 2%, during transfection, seems to protect cells better, without reduction in transfection rate. good luck.
Replace medium containing transfection mixture after 5 hrs. Add a low % FBS, 2%, during transfection, seems to protect cells better, without reduction in transfection rate. good luck.
I'll try that and see. Thanks