The luciferase gene or luciferase bioluminescence activity does not affect the growth of cancer cell lines in vitro. (A) MCF-7 cells were assessed for proliferation by MTT assay: i) untransduced; ii) transduced with GFP alone; iii-v) transduced with GFP-P2A-luc (clones 1-3); vi) representative flow cytometric analysis of GFP expression in cells i-v. (B) B16-F10 cells were assessed for proliferation by MTT assay: i) untransduced; ii) transduced with GFP alone; iii-vi) transduced with GFP-P2A-luc (clones 1-4). Cells were treated with D-luciferin substrate (150 μg/mL final concentration) diluted in media or media alone at 2-day intervals. Following 30 min incubation at 37°C in the dark, cells were washed once with PBS and the normal growth medium was replaced. Proliferation was measured by addition of MTT substrate overnight, followed by measuring the absorbance at 572 nm. Data is shown as the mean ± s.e.m. from 3 independent experiments.