I'm trying to evaluate the effect of a mutation in the promoter region on translation using dual luciferase assay. I transfected HL60 cell line using 4 ug of pGL4.10 [luc2] and 444ng of pGL4.74 [HRL-TK] as a reporter. In this way I was able to obtain a transfection efficiency of  70% with Nucleofector (Amaxa). I repeated the experiments for 5 times, each samples in triplicate. Triplicate were very reproducible  in the same experiment. The big problem is that data are not reproducible between different experiments. It might depend on the cellular system I'm using? HL60 cells do not survive for more than 24 hours after transfection with the Nucleofector. I harvested cells after 20 hours from transfection to avoid this problem. More over relative luciferase units of renilla between different samples are extremely variable. May be it a problem? 

Thank you in advance,