Figure 2.

ELISPOT analysis of CD8⁺ T cells from PBMCs demonstrates postvaccination induction of mesothelin-specific T cells in three DTH responders. (A) ELISPOT analysis of PBLs from two patients who were HLA-A3⁺. (B) ELISPOT analysis of PBLs from two patients who were HLA-A-2 and HLA-A3⁺. (C) ELISPOT analysis of PBLs from two patients who were HLA-A24⁺. (D) ELISPOT analysis of PBLs from all 14 patients who were treated with the vaccine (reference 24). ELISPOT analysis for IFN-γ–expressing cells was performed using PBMCs that were isolated on the day before vaccination or 28 d after the first vaccination. T2-A3 cells were pulsed with the two mesothelin-derived epitopes MesoA3_(83–91) (squares), MesoA3_(225–233) (X), and HIV-NEF_(94–102) (not shown). T2-A2 cells were pulsed with the two mesothelin-derived epitopes MesoA2_(20–28) (triangles), MesoA2_(530–538) (circles), and HIV-gag_(77–85) (not shown). T2-A24 cells were pulsed with the two mesothelin-derived epitopes MesoA24_(435–443) (asterisks), MesoA24 _(475–483) (diamonds), and tyrosinase A24_(206–214) (not shown). All DTH responders are represented by dotted lines and open symbols, and DTH nonresponders are represented by solid lines and closed symbols. For the detection of nonspecific background, the number of IFN-γ spots for CD8⁺ T cells specific for the irrelevant control peptides were counted. The HLA-A2 binding HIV-gag protein-derived epitope (SLYNTVATL), the HLA-A3 binding HIV-NEF protein-derived epitope (QVPLRPMTYK), and the HLA-A24 binding tyrosinase protein-derived epitope (AFLPWHRLF) were used as negative control peptides in these assays. Background was minimal to negative control peptides, ranging from zero to three spots total (data included in each graph). Data represent the mean of each condition assayed in triplicate, and standard deviations were <5%. The number of human IFN-γ spots per 10⁵ CD8⁺ T cells is plotted. Analysis of each patient's PBLs was performed at least twice.