Sphere formation assay

Sphere formation assay was performed based on the previously described method [31]. The ES-2 and OVCAR-3 cells were plated at different oxygen tensions for 48 hrs, and then dispatched from cell culture flask and harvested. Single cells (1000 cells per well) were re-plated and cultivated under normoxia at ultralow attachment six-well plates (ultra low cluster plates, Life sciences). These cells were cultivated for 14 days under normoxic condition before the spheres were evaluated under inverse miscopy and counted (more than 30 cells within a sphere was considered to be a full sphere). In addition, sphere formation assays of single CD44^bright and CD44^dim cells (800 cells per well) were also performed with the method as described above. All the experiments were repeated three times.