This study demonstrates that gene-gun inoculation of mice with DNA encoding the influenza virus hemagglutinin (HA) results in the life-long maintenance of protective B cell responses. Using a sensitive single-cell enzyme-linked immunospot assay, we show that all of the HA-specific plasma cells are localized in the bone marrow and spleen 1 year postimmunization. As a consequence of prior virus challenge, only a small population of antibody-forming cells was found in the lymphoid tissues associated with the respiratory tract. The tissue distribution of HA-specific plasma cells in these mice was identical to the profile in infected controls. Complete protection against live virus challenge in the aged vaccinated mice did not require prior exposure to virus. Thus, immunization with the DNA, vaccine provides long-term protective immunity against otherwise lethal infection.