Aims: To isolate new lactobacilli strain with cholesterol-lowering effect and analyse its probiotic properties and possible mechanisms of cholesterol removal. Methods and Results: The strain with cholesterol-lowering effect was isolated from pickled juice. The acid and bile tolerance and antimicrobial activity were tested. The free cholalic acid, the cholesterol in supernatant fluid, washing buffer and cell extract, the cholesterol removed by growing, dead and resting cells were quantified. The isolated strain with high cholesterol-reducing rate of 43 center dot 95% was identified as Lactobacillus buchneri (Lact. buchneri) P2. It had acid and bile tolerance and antimicrobial activity. Moreover, it could remove cholesterol via coprecipitating with deconjugated bile salts, assimilating and adsorbing by cells. And the assimilation was considered to be the main reason of cholesterol removal. Conclusions: The isolated Lact. buchneri P2 showed probiotic properties of cholesterol reduction, acid and bile tolerance and antimicrobial activity and could remove cholesterol via different ways. Significance and Impact of the Study: A new strain of Lact. buchneri P2 with efficient cholesterol-reducing ability was isolated to provide species diversity of lactobacilli for functional dairy products. And the possible mechanism of cholesterol removal by Lact. buchneri was discussed.