Chitosan was used for nanoencapsulation of Lactobacillus acidophilus as probiotic bacteria. In vitro experiments were done with the objective of investigating the survival of the bacteria cells in gastro-intestinal conditions. The results demonstrated that the size of chitosan nanoparticles noticeably increases by increasing chitosan concentration from 0.05 to 0.5 g/mL. Encapsulation of the cells caused a decrease at the leakage of probiotic bacteria when compared with free bacteria. However, the number of probiotic cells reduced from 3.27 to 3.23 log CFU/ml in gestural acid condition in contrast to free cells which they approximately dropped from 3.3 to 3 log CFU/ml after 120 min. Good probiotic viability and stability was also obtained by nanocapsulation of Lactobacillus acidophilus in intestinal juice. At this condition, the initial capsulated cells numbers were 3.28 log CFU/ml which after placing in biliary salt condition for 120 min, it reached to 3.23 log CFU/ml. On the other hand, the free bacteria cells reduced from 3.3 to 2.97 log CFU/ml in intestinal environment. Overall, nanocapsulation of probiotic bacteria plays a pivotal role in enhancing the viability and survival of them against gastro- intestinal environmental conditions.