In order to determine which formulation was most immunogenic, vaccination with a single peptide or a mixture of the peptides was compared with linked lipopeptide vaccines. Results of a representative experiment are presented in Figure 3 . Mice immunized with a vaccine formulated with a single peptide SAG1 224-232 , KS9-PADRE with the adjuvancy of GLA-SE and Pam 2 Cys, elicited IFN-γ production (SFC:248 ± 65; mean ± SEM) four-fold higher (p < 0.01) than mice that received LpKS9 GLA-SE vaccination (63 ± 17) (Figure 3a ). Similar results, with substantial IFN-γ production, were found when splenocytes from mice immunized with a mixture of peptides and adjuvants. These results were compared with IFN-γ production by splenocytes from mice immunized with a lipopeptide vaccine constructed with the three peptides linked together with alanine spacers LpKS9-AM9-RS9. Significant IFN-γ production was only present with KS9 and AM9 stimulation of spleen cells, and was much less robust and not significant when the spleen cells were stimulated by RS9 peptide.