FIG 3.

Immunization with either ID83/GLA-SE or ID93/GLA-SE stimulates pluripotent Th1 antigen (Ag)-specific responses. Mice were injected s.c. with ID83/GLA-SE and ID93/GLA-SE at biweekly intervals for a total of 3 immunizations. Single-cell suspensions of spleen cells were prepared 1 month after the final immunization and cultured with 10 μg/ml protein. Culture supernatants were collected after 4 days, and IFN-γ (A) or IL-5 (B) content was determined by ELISA. Results are shown as means and SE; n = 3 per group. iono, ionomycin. Alternatively, cells were cultured with antigen and BD Golgi Stop for 16 h and then fixed and stained to determine the percentage (C) of CD3⁺ CD4⁺ CD44^hi IFN-γ⁺ cells by flow cytometry. Results are shown as means and SE; n = 3 per group. The phenotype of each CD3⁺ CD4⁺ CD44^hi IFN-γ⁺ cell was further delineated by costaining for IL-2 and/or TNF. In panel D, results are shown as percent CD3⁺ CD4⁺ CD44^hi IFN-γ⁺ cells exhibiting each phenotype. Data are representative of at least 3 independent experiments.