Abstract: The inhibitory effect of lactic acid bacteria on the pathogenic bacteria of dairy cow mastitis was studied to reduce the economic loss caused by bovine mastitis. In this experiment,Escherichia coli ATCC25922,Staphylococcus aureus ATCC25923 and Streptococcus agalactiae ATCC14556 were used as indicator bacteria. A strain with better bacteriostatic was selected from 8 strains of Lactobacillus by using antibacterial test and adhesion test,and the antibacterial mechanism was analyzed. The results showed that the inhibitory effect of Lactobacillus KLDS1.0344 on indicator bacteria was more obvious than that of other Lactobacillus. The maximum inhibitory diameter was 15.4 mm. KLDS1.0344 was Lactobacillus plantarum indentified by 16S rDNA. Preliminary studies on the antibacterial mechanism indicated that the lactic acid and bacteriocin substances produced in the metabolic process played a major role in inhibiting the three indicator bacteria. These antibacterial active substances had stronger antibacterial activity under acidic conditions.