Summary A hemagglutinin fraction (HAF) of influenza A (PR8) virus, purified by sucrose density gradient centrifugation and shown to be highly homogenous by analytical ultracentrifugation, electron microscopy and a gel diffusion test, was used as an immunizing antigen in these studies. Sera of guinea pigs immunized intraperitoneally with purified HAF contained antibodies which exhibited hemagglutination-inhibiting, virus neutralizing, and toxin neutralizing properties. Mice immunized with purified HAF prepared from PR8 virus were resistant to challenge infection with the homologous strain of virus but susceptible to infection with a heterologous (FM1) strain of virus. Guinea pigs immunized with whole virus and challenged intranasally with homologous virus exhibited more extensive and intensive histopathologic changes than nonimmunized animals; intranasal challenge of immunized animals with purified HAF duplicated some of the pathologic changes caused by challenge of immunized animals with whole virus.