To optimize the luciferase light reaction, we tested a series of flash and glow luminescent substrate systems in both lysed and live 2fTGH-ISRE-CBG99 and HEK293T-ISRE-CBG99 cells, including D-luciferin (Fisher Scientific, Pittsburgh, PA), the Chroma-Glo Luciferase assay system from Promega and the steadylite plus reporter gene assay system from Perkin Elmer (Waltham, MA) under a range of incubation conditions. The steadylite plus system was selected based on kinetic profile. Effects of IFN-b concentra-tion and treatment time were assessed with all other variables constant.