Hyclone cell molecular biology grade

A TP ODN stock solution in water (Hyclone Cell Molecular biology Grade, Invitrogen) was added to 375 μl reduced serum without antibiotic medium (2% FBS, 1× GlutaMAX, 1× non-essential amino acids and 1× sodium pyruvate) so as to obtain a final concentration of 1 μM TP ODN. Medium was removed from 15b HeLa cells in a 96-well plate and 100 μl of the transfection solution was added to each well. After 72 h of passive transfection, cells were collected for luciferase assays.

To 50 μl of a reduced serum, no antibiotic medium (2% FBS, 1× GlutaMAX, 1× non-essential amino acids and 1× sodium pyruvate) was added a stock solution of a TP ODN in water (Hyclone Cell Molecular biology Grade, Invitrogen) so as to obtain a final concentration of 0.1 μM TP ODN. In an eppendorf tube, 0.25 μl DharmaFECT 1 (purchased from Dharmacon/ThermoFisher, now GE Dharmacon, Lafayette, CO, USA) was added to 50 μl of the reduced serum medium and this solution was added to the TP ODN in the same medium. TP ODN and DharmaFECT 1 (Dharmacon) were mixed and allowed to form a complex at room temperature (20 min). A measure of 275 μl medium was added to the above complex. Medium was removed from 15b HeLa cells in a 96-well plate and 100 μl of the transfection mixture was added to each well. After 24 h of lipid transfection, cells were collected for luciferase assays.