All antigens were >95% pure as judged by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and staining with Coomassie brilliant blue R250 (37)

All antigens were >95% pure as judged by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and staining with Coomassie brilliant blue R250 (37). The PdB antigen was a genetic toxoid of pneumolysin which has a Trp433Phe mutation reducing cytotoxicity and retains full immunogenicity (41). than those of ASCs for PspA and PdB (pneumolysin toxoid B) (< 0.001). For all those antigens, the numbers of IgA PU 02 ASCs tended to be lower than those of both IgG and IgM ASCs. The numbers of anti-CbpA and -PsaA IgA ASCs were higher than those of anti-PdB IgA ASCs (< 0.01). Concentrations of IgA antibodies to PspA and PsaA in saliva correlated with the numbers of IgA ASCs to PspA and PsaA in freshly isolated adenoidal cells, but no such correlation was found between salivary IgG antibody concentrations and IgG ASCs to the four antigens in adenoidal cells. In cultured cells, anti-PspA, -PsaA, and -CbpA IgG ASCs proliferated significantly, but only two of eight samples showed >2-fold increases in anti-CbpA and -PspA IgA ASCs after CCS stimulation. The results suggest that CbpA, PsaA, and PspA may be good upper respiratory mucosal antigens in children. Adenoids may be important inductive sites for memory IgG responses and important sources of salivary IgA. Some protein antigens may also primary for mucosal IgA memory. These data support the effort to explore mucosal immunization against pneumococcal contamination. is usually a common cause of otitis media, pneumonia, septicemia, and meningitis in children, resulting in significant mortality and morbidity throughout the world. With the prevalence of antibiotic-resistant pneumococci increasing worldwide (20, 26), studies of pneumococcal vaccines have gained much interest. The efficacy of polysaccharide vaccines is limited by poor immunogenicity in high-risk populations, especially young children. Conjugate vaccines have greater immunogenicity than polysaccharide-based vaccines, but serotype coverage is limited. Efforts are being made to find effective pneumococcal protein vaccines which might protect against multiple serotypes and which are immunogenic in children as well as in adults. Currently, several candidate pneumococcal proteins are under study, including pneumolysin, pneumococcal surface protein A (PspA), pneumococcal surface adhesin A (PsaA), and choline-binding protein A (CbpA; also referred to as PspC, SpsA, or Hic) (11, 19, 21, 44). Of these pneumococcal protein antigens, pneumolysin, PspA, and PsaA have been shown to contribute to the virulence of pneumococci and to be produced by virtually PU 02 all clinical isolates (35, 39). CbpA is likely to play a role in nasopharyngeal colonization and appears to be expressed by most, if not all, isolates. Preliminary studies of mice have shown Smad1 that immunization with these proteins can protect against contamination with multiple serotypes of pneumococcus (1) and/or prevent nasopharyngeal carriage (9, 10). It has been shown that pneumococcal carriage and infections induce salivary and serum antibodies to PsaA, pneumolysin, and PspA in children (43, 46, 48). As pneumococci are mucosal pathogens colonizing the nasopharynx, mucosal immunization (e.g., by the intranasal route) is potentially a better way to protect against mucosal carriage than parenteral immunization. Acquisition of pneumococci is generally from nasopharyngeal carriers rather than infected individuals. Therefore, to induce community immunity against gains entry into the host via the epithelium of the upper respiratory tract. Asymptomatic carriage of pneumococci is particularly common in infants and young children (17, 52), age groups also at high risk of invasive disease. Previous studies suggest that natural mucosal infections (or carriage) can be immunizing processes which can primary tonsillar lymphocytes. Natural contamination or intranasal immunization with rubella virus vaccine primes tonsillar lymphocytes better than subcutaneous vaccination (34). Natural contamination with varicella-zoster virus likewise stimulates tonsillar lymphocytes better than peripheral blood lymphocytes (4). Adenoids (nasopharyngeal tonsils), which are located in the anatomical area of pneumococcal carriage, are thought to be important immune inductive and effector PU 02 sites for nasopharyngeal immunity and to act as a part of an integrated mucosal immune system (25). Regional mucosal immunity induced by natural carriage or intranasal vaccination most probably involves PU 02 these immunocompetent tissues in the nasopharynx. Thus, protein-based vaccines against pneumococci should ideally be immunogenic to adenoidal lymphocytes in children if they are candidates for mucosal immunization. Adenoids PU 02 are rich in lymphocytes, especially B cells, and thus provide a good model to study antigen-specific B-cell responses. Adenoidal lymphocytes have been used in a previous study to investigate the immune responses to the P6 outer membrane protein of nontypeable in children (23). Specific immunoglobulin-secreting cells.