The immune response to pneumococcal polysaccharide type 14 (PPS-14), induced by native PPS-14 was compared with the response induced by PPS-14 conjugated with CRM197 (PPS-14-CRM197). In our animal model, immunization with PPS-14-CRM197 gave a significant enhancement of anti-PPS-14 serum titres for IgM and IgG, but not for IgA. Also an increase in total number of anti-PPS-14 antibody-secreting cells was found. Using immunohistochemical techniques, a different distribution pattern of specific antibody-containing cells in spleen section after immunization with PPS-14-CRM197 was observed. Furthermore, a higher number of IFN-gamma producing cells was found after immunization with PPS-14-CRM197, as compared with immunization with PPS-14. This enhanced IFN-gamma production may be the cause for enhanced IgG response observed after immunization with PPS-14-CRM197. The specific immune response was less affected by splenectomy in animals immunized with PPS-14-CRM197 than with PPS-14. However, an age-related response to the native as well as the conjugated form of the PPS-14 was observed, since no effect of conjugation with CRM197 was seen in the onset of the immune response to PPS-14 in young animals. In conclusion, our results affirm the hypothesis that conjugation of polysaccharides changes the characteristics of the antigen towards a thymus-dependent antigen.