Use of efficacious vaccines remains one of the most cost effective methods to prevent disease. New vaccine delivery vehicles are needed to improve effectiveness and patient compliance. Biodegradable polyanhydrides have shown great promise as single dose vaccine delivery vehicles that also function as superb adjuvants. Our current studies are designed to test polyanhydride nanosphere uptake and intracellular localization in antigen presenting cells since these are key determinants in antigen presentation. Copolymers of sebacic acid and 1,6-bis(p-carboxyphenoxy)hexane and copolymers of 1,8-bis(p-carboxyphenoxy)-3,6-dioxaoctane and 1,6-bis(p-carboxyphenoxy)hexane were tested as potential vaccine adjuvants. Polyanhydride nanospheres were fabricated using the nanosphere antisolvent precipitation technique. Murine bone marrow derived dendritic cells were incubated in vitro with nanospheres and analyzed for increases in cell surface marker expression by flow cytometry. Internalization and intracellular distribution of fluorescein-loaded nanospheres within human monocytic cells were visualized using laser scanning confocal microscopy. It was observed that internalization and particle trafficking were strongly dependent on specific copolymer chemistry and particle size. These studies provide evidence that polyanhydride nanospheres are promising candidates for vaccine delivery. In addition, the effect of size and chemistry can be exploited to design delivery systems that facilitate appropriate and efficacious immune responses.