The replicon of Semliki Forest virus (SFV) offers the possibility to direct high-level, transient expression of heterologous proteins in vivo. We initiated studies to determine the possibility of employing the SFV expression system for recombinant vaccine purposes. Mice immunized with recombinant SFV encoding Influenza A nucleoprotein (NP) or E. coli LacZ developed long-lasting antigen-specific IgG levels and induction of cytotoxic T-cell (CTL) memory that persisted for over one year. Predominantly type 1 T-helper cells were induced as shown by IgG subclass ELISA. Humoral and cell-mediated immune responses could be induced upon delivery by several administration routes and mucosal immunizations induced secretory IgA in the respiratory tract. Development of immune responses against the vector itself did not inhibit boost responses by subsequent immunizations with recombinant SFV. Immunization of mice with vectors encoding the Influenza A virus antigens nucleoprotein (NP) and hemagglutinin (HA) resulted in immune responses that were protective against challenge infection with Influenza virus.