After incubation with standard and unknown samples, bound immunoglobulins were detected with peroxidase-conjugated goat antibodies to human IgA, IgG, or IgM (Sigma). The pattern of serum antibody specificities against the different OMV components after intranasal immunizations was largely similar to that obtained with the intramuscular vaccine. Five and eight vaccinees in the nasal group developed persistent increases in serum bactericidal titers to the homologous meningococcal vaccine Vasopressin antagonist 1867 strain expressing low and high levels, respectively, of the outer membrane protein Opc. Our results indicate that meningococcal OMVs possess the structures necessary to initiate systemic as well as local mucosal immune responses when presented as a nasal vaccine. Although the serum antibody levels were less conspicuous than those after intramuscular vaccinations, the demonstration of substantial bactericidal activity indicates that a nonproliferating nasal vaccine might induce antibodies of high functional quality. Vaccines administered directly onto mucosal surfaces may induce local mucosal as well as systemic immune responses (23,24). Even nonproliferating mucosal vaccines may thus offer a challenging alternative to traditional parenteral vaccines, as has been shown for an oral cholera vaccine (15). It is required, however, that induction of tolerance to antigenic components of such vaccines be abrogated or that so-called mucosal PCK1 adjuvants be added (10,24). We have shown that in mice, the nasal mucosa is the preferred site for presentation of a vaccine consisting of whole killed pneumococci in suspension, with cholera toxin (CT) added as mucosal adjuvant (1). Even for intestinal immune responses, as measured by antibodies in feces, nasal immunizations were superior to administering the antigen by both the oral and gastric routes. Subsequently, we found that outer membrane vesicles (OMVs) from group B meningococci were also immunogenic in mice when given nasally (8). The antibody responses to OMVs in these experiments were largely independent of adding CT; i.e., the vesicles themselves possessed the necessary structures for induction of mucosal and systemic immune responses after application on mucosal surfaces. Outer membrane proteins from group B meningococci are clearly immunogenic in humans (31), and the OMVs which we used as a mucosal vaccine in mice were originally developed to be the main component of a parenteral vaccine against group B meningococcal disease (12). In a large-scale study of adolescents, this OMV vaccine was shown to protect against disease when given intramuscularly with aluminum hydroxide as adjuvant (6). In the present study, we used OMVs, suspended in saline without aluminum hydroxide, as a mucosal vaccine in the form of nasal drops or spray to human volunteers. The demonstration by others of M cells in the human nasopharyngeal area (30) forms the basis for an effect of such a vaccine when applied intranasally (19). Other researchers have recently also demonstrated that intranasal immunizations with either live influenza virus (18), the B subunit of CT (CTB) (4), or diphtheria-tetanus vaccines (2) can induce specific immune responses in humans. The results with our nasal OMV vaccine against meningococcal Vasopressin antagonist 1867 disease were compared with those obtained in another Vasopressin antagonist 1867 group of volunteers who received two intramuscular doses of the parenteral OMV vaccine formulation with aluminum hydroxide. The aim was to determine whether intranasal delivery of such particles might also induce immune responses in humans, and that they might serve as a model system for creating alternative mucosal vaccines against other bacterial diseases. == MATERIALS AND METHODS == == Vaccinees. == Twelve healthy volunteers, nine women and three men at 25 to 61 (median, 46) years of age, were included in the nasal vaccine study regardless of their prevaccination antibody levels. They had not previously received a meningococcal vaccine and did not receive other vaccines during the study. Another group of 11 healthy volunteers, seven women and four men at 24 to 49 (median, 38) years of age, were immunized intramuscularly with the regular vaccine formulation and served as controls for the nasally immunized volunteers. This group of volunteers was selected on the basis of low serum immunoglobulin G (IgG) antibody levels to meningococcal OMVs. The reason for different selection criteria in the two groups of volunteers is that the study was originally planned as.