Streptococcus pneumoniae is a major pathogen in humans that enters the host primarily through the respiratory tract. Targeting mucosal surfaces directly may therefore be an optimal approach for vaccination to prevent bacterial colonization and invasive disease. We have previously demonstrated the effectiveness of interleukin-12 (IL-12) delivered intransally (i.n.) as an antiviral respiratory adjuvant. In this study, we examined the effects of i.n. IL-12 treatment on induction of protective humoral immunity against S. pneumoniae. Immunization i.n. with pneumococcal surface protein A (PspA) and IL-12 resulted in enhanced lung IL-10 mRNA expression and marked augmentation of respiratory and systemic immunoglobulin G1 (IgG1), IgG2a, and IgA antibody levels compared to those in animals receiving PspA alone. In addition, i.n. vaccination with PspA and IL-12 provided increased protection against nasopharyngeal carriage. Flow cytometric analysis revealed a threefold increase in antibody mediated, complement-independent opsonic activity in the sera of PspA- and IL-12-treated animals, which was mainly contributed by IgG2a and, to a lesser extent, IgA. Passive transfer of these immune sera conferred complete protection from death upon systemic pneumococcal challenge. These findings demonstrate the effectiveness of combining PspA and IL-12 at mucosal sites to achieve optimal antibody-mediated opsonization and killing of S. pneumoniae.