Cell Viability Assay for Radiosensitization

Cells were plated using a Multidrop Combi liquid handler (Thermo Fisher) in at least quadruplicates for each time point at three cell densities (range 25–225 cells per well) in a white 384-well plate (Corning, NY). Plates were irradiated and at 9 days post irradiation, media was aspirated and 40 μl of CellTiter-Glo reagent (50% solution in PBS; Promega, WI) was added to each well. Relative luminescence units were measured using an Envision multilabel plate reader (Perkin Elmer) with a measurement time of 0.1 s. Luminescence signal is proportional to the amount of ATP present. For chemical radiosensitization measurements, drug was added 24 h before irradiation. The luminescence signal was plotted as a function of cell density and a cell density within the linear range for luminescence (or growth) was selected to generate integral survival for each cell line (see also Supplementary Data in ref. 8 (link)).