As shown in Figure 2, proliferation of splenocytes stimulated with the 8-epitope mixture (mix2) was more selleck screening library significant comparing to single-epitope phages or 4-epitope mixture of OmpL1 or LipL41 alone (mix1). Evaluation of cytokine secretion in splenocytes induced by OmpL1- or LipL41-derived epitopes ELISA assay was employed to determine the in vitro polarization of

T helper cells. Cells from both OmpL1- and LipL41-immunized mice released large amount of IFN-γ but not IL-4 comparing to cells from PBS control mice (Figure 3). OmpL1173-191 epitope showed the strongest activity of stimulation, and other three OmpL1 epitopes showed similar abilities in the stimulation of IFN-γ secretion. Among the LipL41 epitopes, the secretion of IFN-γ in the cell cultures was induced by WZB117 LipL41181-195, LipL41233-256 and LipL41263-282 to the similar level; all of them were stronger than LipL4130-48. When the 4 epitopes of OmpL1 were pooled together to stimulate the splenocytes, the secretion of IFN-γ cytokine in the splenocyte supernatants was mildly increased. Phages expressing each epitope of LipL41 failed to stimulate the secretion of IFN-γ or IL-4 (Figure 3B). Figure 3 Cytokine profiles of T cells from mice spleen. Splenocytes from recombinant see more OmpL1 (A) or LipL41

(B) immunized mice were isolated 10 days after the last immunization and were stimulated with epitopes from corresponding proteins in vitro for 72 hours. Mix stand for the data from the epitope mixture of OmpL1 or LipL41 stimulating the splenocytes from OmpL1- or LipL41- immunized mice. Each value is representative of 3 mice in triplicates. Discussion Leptospira interrogans causes disease in both animals and humans throughout the world. Leptospirosis in humans may be fatal due to the involvement of severe damage to multiple organs such as liver, lung, kidney and brain and is an increasing concern to the public health [24].

L. interrogans can rapidly disseminate to multiple organs to induce programmed cell death [25, 26]. The essential properties of a vaccine are safe, immunogenic, and effective in the prevention of leptospiral infection at both acute and carrier many state. It has been a challenge to develop an effective and safe L. interrogans vaccine [27]. The currently available vaccines include multiple-valence inactivated leptospiral vaccine and subunit leptospiral vaccines [28]. However, these vaccines often have serious adverse effects [29]. And more importantly, most recombinant protein vaccines used against Leptospira in animals are serovar-specific and therefore their efficacy is limited when Leptospira of a different serovar is circulating [30]. The current emphasis in research laboratories is to discover conserved antigens that may induce long term protection across the species or serovars of Leptospira.