In this study, we evaluate the effect of phospholipid on the adjuvanicity and protective efficacy of liposome vaccine carriers against visceral leishmaniasis (VL) in a hamster model. Liposomes prepared with distearyol derivative of l-α-phosphatidyl choline (DSPC) having liquid crystalline transition temperature (Tc) 54 C were as efficient as dipalmitoyl (DPPC) (Tc 41 C) and dimyristoyl (DMPC) (Tc 23 C) derivatives in their ability to entrap Leishmania donovani membrane antigens (LAg) and to potentiate strong antigen-specific antibody responses. However, whereas LAg in DPPC and DMPC liposomes stimulated inconsistent delayed type hypersensitivity (DTH) responses, strong DTH was observed with LAg in DSPC liposomes. The heightened adjuvant activity of DSPC liposomes corresponded with 95% protection, with almost no protectivity with LAg in DPPC and DMPC liposomes, 4 mo after challenge with L. donovani. These data demonstrate the superiority of DSPC liposomes for formulation of L. donovani vaccine. In addition, they demonstrate a correlation of humoral and cell-mediated immunity with protection against VL in hamsters.