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Aims:It suggests that cancer is maintained mainly by a few cancer cells with stem cell properties,named as cancer stem cells(CSCs).The isolation and expansion of CSCs is crutial for understanding cancer biology.However,conventional methods is always unreliable or lower-producing.For this reason,we attempt to a new method to enrich cancer stem cell.Methods:We developed a new simple method for selecting and expanding CSCs with cotton filers as a supporting material.Results:In this culturing system,PC12 grew into individual round colonies with strong regenerative ability and drug resistance capability.The western blot analysis showed the expression of the CSC-related proteins(Oct4,Sox2,Nanog)in PC12 spheroid were higher than that of monolayer cells.Conclusion:We developed a new method for selection and enrichment of cancer stem cells.Our study may contribute to obtain mass number of cancer stem cells and support the cancer therapy studies,such as evaluating drug resistance ability of cancer.