The immunogenicity of a mucosally delivered subunit influenza vaccine was assessed in mice. Split influenza virus vaccine (sFlu) was formulated with proteosomes (Pr-sFlu), administered intranasally, and the induced immunity was compared with the responses elicited by sFlu alone given either intramuscularly or intranasally. Intranasal (i.n.) immunization with Pr-sFlu induced specific serum IgG and hemagglutination inhibition (HAI) titers comparable to or better than those induced by intramuscular (i.m.) sFlu, and in contrast to sFlu alone, i.n. Pr-sFlu also induced high levels of influenza-specific IgA in lung and nasal washes. Mice receiving i.n. Pr-sFlu were completely protected against live virus challenge, as were mice immunized by injection with sFlu alone. The i.n. Pr-sFlu elicited cytokine responses polarized towards a type 1 phenotype whereas those elicited by sFlu alone were of a mixed type 1/type 2 phenotype. The data strongly suggest that i.n. proteosome-formulated influenza antigens are highly effective and are excellent candidates for a non-invasive human vaccine.