Reverse Transfection of Plasmid into Glioma Stem Cells Using Lipofectamine 3000 Reagent

In Vial 1 (125uL of opti-MEM was mixed with 7.5 uL of lipofectamine reagent). Another vial 2 (125ul opti-MEM + 3ug of plasmid + 5ul of p3000 reagent) was mixed and incubated for 5 min. Contents from Vial 1 and 2 were mixed well and incubated for 10 min at room temperature, added to 6 well plates. Glioma stem cells were seeded at a density of 1 million on top of transfection contents in 6 well plates (total volume of medium is 2 ml per 6 well plate). 48 hour post-transfection cells were lysed and examined for the expression of expressed proteins by western blot.