Mouse i.m. immunization with MecVax.

A total of 32 8-week-old female BALB/c mice (Charles River Laboratories, Wilmington, MA) in four groups with eight mice per group were included for immunization. One group was intramuscularly (i.m.) immunized with MecVax containing 25 μg CFA/I/II/IV MEFA protein and 25 μg toxoid fusion protein 3xSTa_N12S-mnLT_R192G/L211A in a total volume of 50 μl under a regimen of one primary immunization followed by two boosters at intervals of 2 weeks. The second group and the third group were i.m. immunized with 25 μg CFA/I/II/IV MEFA protein alone or 25 μg toxoid fusion 3xSTa_N12S-mnLT_R192G/L211A protein alone. By comparatively analyzing antigen-specific antibody responses and antibody protective activities in the first group and the second or the third group, we evaluated MecVax protein antigenic compatibility. The adjuvant dmLT (0.2 μg) was used for each immunization group. The fourth group was i.m. immunized with 50 μl phosphate-buffered saline (PBS) and served as the control group.