Figure 5. CD40L-sBAFF-B cells pulsed with TL-9 provided an effective APC source to generate HIV-1-specific CTLs.

At day 5 after co-culturing, CD40L-B cells and CD40L-sBAFF-B cells were harvested and incubated with peptide TL9 (20 ng/ml) at 37°C for 12 hours. Then autologous CD8⁺ T cells were co-cultured at a T: B ratio of 3:1 for 7 days with these irradiated B-cells. On day 12, CD8⁺ T lymphocytes were harvested, washed, and restimulated with fresh TL9-pulsed B cells and IL-2. This was repeated on days 19 and 26. After that, CTLs were co-cultured with HIV-1_{NL4-3□env-EGFP}-infected primary CD4⁺ T cells as target cells. a, b. Representative results of IFN-γ production produced by the educated CTLs. The IFN-γ production was measured by ELISPOT assay. c, d. The educated HIV-1-specific CTLs were mixed with HIV-1_{NL4-□env-EGFP}-infected primary CD4⁺ T cells as target cells at different E: T ratios. The cell lysis rate was measured with LDH-release assay. e. The calculation of survived HIV-1-infected CD4⁺ GFP⁺ cells after co-culture with the educated CD8⁺ T cells. The CD8 T-cells were educated with various B-cells. Then HIV-1-infected CD4⁺ GFP⁺ cells were mixed with the CD8 T-cells at E:T ratio of 3:1 in the presence of IL-2 for 6 days. The proportion of survived GFP⁺ cells was measured with FACS analysis. The survived GFP⁺ cells mixed with the CD8⁺ T-cells generated with TL9 unpulsed CD40L-B cells at day 0 were set as 100%. Data represent mean ± SD (error bars) for a representative experiment of three independent experiments. The student t-test and one-way ANOVA were used. P < 0.05 indicates statistically significance difference. * indicates P < 0.05; ** indicates P < 0.01; *** indicates P < 0.001.