For the evaluation of INF-c positive CD8+ T cells 10 days postimmunization, splenocytes were harvested and cultured (1610 6 / sample) for 5 hours at 37uC in 96-well round bottom microtiter plates in DMEM supplemented with 10% FBS, 2-beta-mercaptoethanol (10 26 M) and GolgiStop (1 ml/ml, BD PharMingen, San Diego, CA). The TELRTFSI peptide which carries the Ebola Zaire GP immunodominant MHC class I epitope for mice of the H-2 k haplotype (B10.BR) was used for stimulation at a concentration of 1 mg/ml [42] . Control cells were treated with either an unrelated peptide or no peptide. After washing, cells were stained with 100 ml of a FITC-anti mouse CD8a antibody (1:100 dilution, PharMingen) at 4uC for 30 minutes. Cells were washed again, permeabilized in 16Cytofix/Cytoperm (PharMingen) for 20 minutes at 4uC, washed with 16Perm/Wash (PharMingen) and stained with 100 ml of a PEanti mouse IFN-c antibody (1:100 dilution, PharMingen) in the same buffer at 4uC for 30 minutes.