Biodegradable nanoparticles with surface adsorbed antigens represent a promising method for in vivo delivery of vaccines targeting a wide range of infectious diseases or cancers. We investigated the feasibility of loading dendritic cells with a vaccine antigen, HIV p24 protein, on the surface of surfactant-free anionic (d,l-lactic acid, PLA) nanoparticles. The p24 protein had a high affinity for the nanoparticles and the antigenicity and immunogenicity of the p24 protein on the nanoparticle was well preserved after immunization. p24-coated nanoparticles were efficiently taken up by mouse dendritic cells (DCs), inducing DC maturation by increasing MHC-I, MHC-II, CD40, CD80 and CD86 surface expression and secreting IL-12 (p70) and IL-4. We evaluated the ability of DCs pulsed with p24-coated nanoparticles to elicit an optimal humoral and cellular immune response in the blood and intestine. DCs pulsed with p24-nanoparticles induced high seric and mucosal antibody production and elicited strong systemic and local lymproliferative responses, correlated with a Th1/Th2-type response, and systemic CTL responses in mice. Thus, DCs pulsed with antigen-loaded PLA nanoparticles may provide a novel delivery tool for cell therapy vaccination against chronic infectious diseases.