Introduction & Objective: Both CD4+ type 1 helper (Th₁) cells and CD8⁺ T cells play effective roles in protection against Mycobacterium tuberculosis infection. DNA vaccine encoding MPB51 can induce Th1-type immune responses and protective immunity upon challenge with M.tuberculosis. This study address to identify T-cell immunodominant epitopes on MPB51 in C57BL/6 mice.

Materials & Methods : We cloned DNA encoding MPB51 molecule in pCI plasmid. After constructing MPB51 DNA-covered gold cartridge, C57BL/6 mice were immunized by using a gene gun system. Two weeks after the last immunization, the immune spleen cells were cultured in the presence of a synthetic overlapping library peptides covering the mature MPB51 sequence or medium alone. Intracellular and cell culture supernatant gamma interferon (IFN-g) production was analyzed using flow cytometry and ELISA, respectively.

Results : Mapping of T-cell epitopes on MPB51 molecule was performed in the spleen lymphocytes restimulated by 20-mer overlapping synthetic peptides of mature MPB51 sequence. Flow cytometric analysis with intracellular IFN-g and the T-cell phenotype revealed that P171-190 and P191-210 peptides contain immunodominant CD4⁺ T-cell epitopes. Further analysis by using T-cell subset depletion and serial peptide dilution revealed that P171 and p191 are H2-A^b-restricted dominant and subdominant CD4⁺ T cell epitopes, respectively.

Conclusion: This study proved that vaccination with plasmid DNA encoding M. tuberculosis-secreted MPB51 protein not only induce CD4⁺ T cells immune response but also is an appropriate method for identifying immunogenic peptides.