In our study, the obtained bovine mammary epithelial cells, which had staining positive for both monoclonal anti-cytokeratin 18 and anti-vimentin, exhibited the same phenomenon as reported previously. The positive staining for cytokeratin-18 was a powerful result to prove the specific epithelium character of CMECs. The phenomenon of CMECs to vimentin was possibly associated with the culture conditions, plastic dish, media, monolayer overspread, and growth without the presence of other cells. The major milk protein genes are defined as mammary-specific and developmentally-regulated expressed genes. As such, they represent markers of mammary differentiation. Epithelial differentiation is characterized by expression of milk proteins, such as b-casein and whey acidic protein, the production of milk fats rich in triglycerides, sources of energy, and essential fatty acids. Casein secretion is the hallmark of the bovine mammary epithelial cells. Acetyl-coa carboxylase plays a pivotal role in the regulation of fatty acid metabolism, which mediates the first committed step for incorporation of acetate carbon into FA. Thereinto, ACACA, a cytosolic protein, provides cytoplasmic malonyl-CoA for FA synthesis, which is rate-limiting for the synthesis of long-chain fatty acids de novo. The enzyme is active in the lactating mammary gland and its activity level is affected by dietary and Triacetin hormonal states of the animal. Butyrophilin, a major milk-fat-globule transmembrane glycoprotein, is also a mammary-specific protein in milk-fat secretion expressed on the apical Testosterone isocaproate surface of the mammary epithelial cells in the final stage of pregnancy and during lactation. And its mRNA could not be detected in bovine heart, intestine, kidney, liver, ovary, or uterus. Although the function of BTN1A1 is not fully understood, its expression profile suggests an important role in lactation. There was no marked difference of ACACA transcript level between mammary gland tissue, isolated epithelial cell cultured in induction media, or resuscitated epithelial cells. However, the transcript level in isolated epithelial cells cultured in basal media was evidently lower.