To investigate the effect of lactobacilli on the Th1/Th2 cytokine and antibody production in sensitized-mice spleen lymphocytes in vitro, BALB/c mice were intraperitoneally injected with β-lactoglobulin, After successfully established the allergic model, the sensitized splenic cells were stimulated with active/heat-killed lactobacilli (107 CFU/mL) at different species in vitro. Supernatants were collected to measure the cytokines (IL-12, IFN-γ, IL-4) and Igs (total IgE, β-Lg-specific IgE, total IgG) production by ELISA assay. In vitro studies, 4 active/heat-killed lactobacilli could differently modulate the production of Th1/Th2 cytokine and antibody, especially the heat-killed Lactobacillus fermentum and Lactobacillus acidophilus were not inferior to the live strains. These two heat-killed Lactobacillus suppressed IL-4 and IL-10 production by stimulating IL-12 and IFN-γ production. Their IFN-γ/IL-4 value, which represents the Th1/Th2 cell balance were higher than that of other strains, and compared with the control were significant (P<0.05). At the same time, these two heat-killed strains significantly decreased the secretion of total IgE, β-Lg-specific IgE and total IgG (P<0.05). These data indicate that lactobacilli may be effective in decreasing the antibody production in sensitized-mice splenocytes by increasing the ratio of IFN-γ/IL-4 and changing the dominant of Th2. Four strains may differ in their respective ability to improve Th1/Th2 cell balance toward Th1 dominance.