New generation vaccines increasingly utilize highly purified peptides and proteins as the target antigen, however these are often poorly immunogenic. One of the most promising strategies for improving immunogenicity of such subunit vaccines is through incorporation into particulate carriers. Here we report the preparation, physicochemical characterization and in vivo immunological activity of cubosomes, a novel lipid-based nanostructured particulate carrier, modified to include the Toll-like receptor agonists monophosphoryl lipid A and imiquimod. The immunological activity of cubosome formulations was compared to that of liposome and alum formulations. Sustained release of the model antigen ovalbumin (Ova) was observed in vitro and in vivo from cubosomes. Cubosomes + adjuvants induced robust CD8+ and CD4+ T cell proliferation and interferon-? production, as well as the production of Ova-specific antibodies. Cubosomes + adjuvants were more efficient at generating Ova-specific cellular responses and were equally as effective in generating humoral responses when compared to liposomes + adjuvants and alum. Overall, the results show that cubosomes have the potential to act as effective sustained release vaccine delivery systems.