Based on the in vitro culturing system developed for epithelial cells in mammary gland of Xinong Saanen dairy goats using tissue explants culture, high density cultivation, and continuous passaging, the cultured epithelial cells were evaluated by growth curve fitting, karyotype analysis, immunofluorescence staining (keratin, epithelial membrane antigen (EMA), vimentin, β-casein), oil red staining and RT-PCR of β-casein gene. The results showed that the growth of epithelial cells with the model number of chromosome of 60 demonstrated a typical ‘S’ shape curve, the positive gene expression of keratin, EMA, vimentin and β-casein was detected, the cytoplasmic lipid droplets were observed following the oil red staining, the cultured cells expressed the mRNA of β-casein. In conclusion, the current in vitro culturing system can obtain the normal mammary gland epithelial cells with the function of secretion.