Objective:To isolate, purify, culture and identify the carcinoma-associated fibroblasts(CAFs) from the human epithelial ovarian neoplasm. Methods:The primary ovarian CAFs were obtained from human epithelial ovarian neoplasm and purified by trypsin digestion and adherence repeatedly. The cells were verified according to morphological observation and immunocytochemical staining of certain proteins. Additionally, their proliferation indexes were assayed. Results:The purified ovarian CAFs were obtained successfully. The typical morphology of ovarian CAFs was simple fusiform fiber like cells. The ovarian CAFs exhibited lower proliferation than tumor cells and showed positive staining for fibroblast specific protein-1, vimentin, desmin and especially for α-smooth muscle actin, and negative staining for cytokeratin(broad-spectrum), cytokeratin 7, cytokeratin 20, CD31 and CD90. Conclusion:The primary cultured CAFs derived from the human ovarian carcinoma tissue can be obtained successfully with digestion method. This provides us an experimental foundation for further studies on the roles of CAFs in the initiation and progression of the cancer.