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One emerging model for the development of drugresistant tumors utilizes a pool of self-renewing malignant progenitors known as cancer stem cells(CSCs)or cancerinitiating cells(CICs).The purpose of this study was to propagate such CICs from the ovarian cancer cell line SKOV3.The SKOV3 sphere cells were selected using 40.0 μmol/l cisplatin and 10.0 μmol/l paclitaxel in serumfree culture system supplemented with epidermal growth factor,basic fibroblast growth factor,leukemia inhibitory factor,and insufin or standard serum-containing system.These cells formed non-adherent spheres under drug selection(cisplatin and paclitaxel)and serum-free culture system.The selected sphere cells are more resistant to cisplatin,paclitaxel,adriamycin,and methotrexate.Importantly,the sphere cells have the properties of se lfrenewal,with high expression of the stem cell genes Nanog,Oct4,sox2,nestin,ABCG2,CD133,and the stem cell factor receptor CD117(c-kit).Consistently,flow cytometric analysis revealed that the sphere cells have a much higher percentage of CD133+/CD117+-positive cells (71%)than differentiated cells(33%).Moreover,the SKOV3 sphere cells are more tumorigenic.Furthermore,cDNA microarray and subsequent ontological analyses revealed that a large proportion of the classified genes were related to angiogenesis,extracellular matrix,integrin-mediated signaling pathway,cell adhesion,and cell proliferation.The subpopulation isolation from the SKOV3 cell line under this culture system offers a suitable in vitro model for studying ovarian CSCs in terms of their survival,self-renewal,and chemoresistance,and for developing therapeutic drugs that specifically interfere with ovarian CSCs.