FIG. 5.

Codon usage effects of DNA vaccine on CTL induction. (A and B) BALB/c mice were immunized intramuscularly with 100 μg of mock vector, pAg85B, or phAg85B twice at 2-week intervals. (C) To compensate for the differences in CpG contents of the pAg85B and phAg85B sequences, mice were injected with a combination of 100 μg each of pAg85B and pGEM-T-hAg85B (pAg85B+phA) or with phAg85B and pGEM-T-Ag85B (phAg85B+pA). Spleen cells from mice immunized with 200 μg of mock vector were used as a negative control. Each of the splenocytes from the immunized mice (5 mice/group) was harvested 2 weeks after the last immunization and stimulated with mitomycin C-treated CT26-Ag85B cells for 6 days in vitro. Cytotoxicity of splenocytes was evaluated in a 4-h ⁵¹Cr-release assay against CT26-Ag85B cells (A and C). Nonspecific lysis was evaluated by using CT26 as target cells (B). The values were expressed as the mean percentage ± standard error of specific lysis from individual mice. The significance of the difference between pAg85B and phAg85B was calculated by a Student's t test. *, P < 0.03; **, P < 0.01.