The TSCC side population was sorted as previously described 29 (link). Briefly, TSCC cells were incubated at 37 °C for 90 minutes in the dark with 5 μg/ml Hoechst 33342 (Sigma, San Louis, MO, USA) and intermittent mixing either alone or in the presence of 50 μM verapamil (Sigma, San Louis, MO, USA), an inhibitor of ABC transporters. The cells were then centrifuged and resuspended in ice-cold HBSS. The samples were sorted using a FACSAria II high-speed cell sorter (BD Biosciences, San Diego, CA, USA). Propidium iodide (2 μg/ml) was added to exclude dead cells. Hoechst 33342 was excited with the UV laser at 350 nm, and the fluorescence emission was measured with 405 / BP30 (Hoechst blue) and 570 / BP20 (Hoechst red) optical filters. After sorting, the SP and non-SP cells were used for further experiments.