The condition of Lactobacillus plantarum formulation was optimized by the single factor and response surface methodology. Meanwhile， the conditions of high-density cultivation， collection of cells and fed batch fermentation method were studied. The results showed that the optimized medium was 2.71% of glucose， yeast 0.45% and beef extract 0.6%. The optimal cultivation conditions were as follows： initial pH 6， inoculation amount 4%， shaking speed 80 r/min， cultivation temperature 35 ℃. Under the conditions of 4 000 r/min and 20 min， Lactobacillus plantarum collection was at the highest survival rate， reaching 81.09%. In addition， the proper addition of nitrogen source can promote the proliferation of the cells， which could reached 9.28 × 109 CFU/mL.