Abstract: Objective To evaluate dynamically cellular immune response of Mycobacterium tuberculosis subunit vaccine AEC/BC-C02 in mice. Methods Forty-eight female BALB/c mice (6-8 weeks old) were randomly divided into two groups. One group was immunized intramuscularly 3 times with AEC/BC-C02 at an interval of 10 days, and the other with PBS as the control. At 1, 2, 4 and 8 weeks after the last vaccination, spleen lymphocytes were isolated (6 mice per group) to measure the frequency of antigen-specific IFN-γ secreting T cell by ELISPOT. Meanwhile, antigen-specific IFN-γ level was detected by ELISA at 2, 4 and 8 weeks, and lymphoproliferation response was measured by cell counting kit-8 assay at 4 and 8 weeks.   Results (1) At 1, 2, 4 and 8 weeks after the last vaccination, the spot forming cells (SFC) of Ag85B-specific IFN-γ in vaccine group were respectively 168.8±103.5, 205.2±51.0, 206.8±65.3 and 160.0±67.9, and significantly higher than that in PBS group (8.9±6.0, t=3.779, P<0.01; 16.1±18.8, t=8.525, P<0.01; 9.3±4.9, t=7.424, P<0.01 and 7.7±6.6, t=5.473, P<0.01, respectively). ESAT6/CFP10 (EC)-specific IFN-γ SFC in vaccine group were respectively 45.1±18.6, 75.6±39.3, 86.2±50.4 and 54.3±26.3, and significantly higher than that in PBS group(4.5±3.5, t=5.258, P<0.01; 11.7±10.5, t=3.850, P<0.01; 3.8±5.8, t=3.977, P<0.01 and 5.2±4.3, t=4.521, P<0.01, respectively). (2) At 2, 4 and 8 weeks after the last vaccination, IFN-γ release induced by Ag85B peptide pool in vaccine group were (1.27±0.13)ng/ml, (1.76±0.55)ng/ml and (1.44±0.44)ng/ml, respectively, and that induced by EC peptide pool were (0.81±0.33)ng/ml, (0.81±0.69)ng/ml and (0.54±0.29)ng/ml, respectively. The difference between them was statistically significant (t=3.008, P<0.05; t=2.631, P<0.05 and t=10.02, P<0.01, respectively). (3) At 4 and 8 weeks after the last vaccination, stimulation indexes (SI) of spleen lymphocytes induced by Ag85B peptide pool were respectively 1.756±0.339 and 1.936±0.287 in vaccine group and signi-ficantly higher than PBS group (1.287±0.0581, t=3.030, P<0.05 and 1.382±0.114, t=4.387, P<0.01, respectively). SI of spleen lymphocytes induced by EC peptide pool in vaccine group were respectively 1.599±0.154 and 1.581±0.156 and significantly higher than PBS group (1.380±0.126, t=2.540, P<0.05 and 1.314±0.170, t=2.844, P<0.05, respectively).  Conclusion Mycobacterium tuberculosis subunit vaccine could induce long term and stable antigen-specific memory T cells in mice, which provided the basis for the late-stage study of its protective immunity against Mycobacterium tuberculosis.

Key words: Vaccines, subunit, Immunity, cellular, Antigens, bacterial, Bacterial proteins, Recombinant fusion proteins, Mice