Abstract: In the mammary glands suffered from invading pathogens that produced and released inflammatory mediators might be accompanied by changes in protein expression profiles. This project presents the protein changes of the mammary gland from healthy dairy cows and clinical mastitic cows that were investigated using two-dimensional gel electrophoresis (2-DE). Acquired gel maps were matched and analyzed using PDQuest 7.4 image analysis software. The results showed that the protein profiles in healthy dairy cows and those with clinical mastitis were shown to be very similar, 480±27 and 453±31 protein spots were statistically detected, respectively. Take "more or less" as the determination criterion, 17 protein spots were detected to be differentially expressed changes in the stain density of threefold or more. That is, 10 protein spots were up or downregulated proteins in clinical mastitic cows, 7 protein spots were unique in healthy dairy cows or clinical mastitic cows. The results indicated significant difference in expression protein between healthy dairy cows and those with clinical mastitis, and may provide valuable information to find new regulation markers of mastitis and potential protein targets for treatment.