Abstract: In this study, a total of 16 LAB strains, isolated from soy sauce residue in the early stage of our research group, were evaluated for their antibacterial activity against Escherichia coli and Staphylococcus aureus. In addition, the bacteriostatic substances and growth characteristics of these strains were investigated. The results showed that LAB strains ZW2, ZW9 and ZW14 exhibited high antibacterial activity against E. coli and S. aureus, of which ZW9 had the best antibacterial effect. After eliminating the interference factors such as the organic acids and hydrogen peroxides, three strains still maintained antimicrobial activity. However, the inhibitory activity decreased by the treatment with trypsin, pepsin, proteinase K, α-amylase, alkaline protease and neutral protease (P<0.05). Therefore, these strains might produce antimicrobial substances of proteinaceous character, i.e., bacteriocins. The three LAB strains showed maximum antibacterial activity and were fairly stable after 20 h of incubation by bacteriostatic profile analysis. The antibacterial substances exhibited a good thermal stability in 40~100 ℃ and kept their high antibacterial activity under ultraviolet irradiation and surfactant treatment. There was no significant decrease in antibacterial activity at pH2.0~6.0, and the antibacterial ability was completely lost at pH10.0. The three LAB strains from soy sauce residue exhibited good antibacterial activity and their antibacterial substances had good stability, which had important reference value for the development of new biological antimicrobial agents.