HCT116/RFP-luciferase cells are derived from the human colon cancer HCT116 cell line by stably integration of a constitutive RFP and Firefly luciferase stably expression construct. The HCT116 cell line has been widely used in cancer research and drug development. HCT116/RFP-luciferase cells stably express RFP and Firefly luciferase, can be used for in vitro assays and in vivo imaging.