Currently, there are three methods that have now been employed to enrich for osteosarcoma CSCs including: (1) the sphere culture assay (or sarcosphere assay), (2) cell sorting for CD133, high ALDH activity, SP cells, or CD117 in combination with Stro-1, and (3) identification of cells that express the embryonic stem cell gene Oct4. This evidence concerns the gene POU5F1 and osteosarcoma.