Objective:To investigate a method of isolating and culturing the epithelial cells from human fistula tube.Methods:We separated epithelial cells of human fistula tube by tissue block method and cultured the epithelial cells.The morphology of the cells was studied with a phase contrast microscope and the living characteristics after subculture of the cells were observed.We identified the cells using immunocytochemical assessment.Results:The culturing cells grew on the wall appeared after 24-36 hours.The living characteristics of the cultured cells were quickly into the logarithms growth period between 7 and 9 days.The identified cells were keratin positive and vimentin negative by using immunocytochemistry.Conclusion:The epithelial cells of human fistula tube could be cultured stably in vitro by the method.It could be possible to study the healing mechanism of human fistula tube based on the results of this study.