We previously reported that a Streptococcus mutans enriched- glucosytransferase (E-GTF) preparation induces an immune response following intranasal, but not tonsillar, immunization of humans. In this study, we determined whether intranasal immunization of these subjects 2 years later resulted in augmented immune responses compared to those seen in control subjects. Subjects previously immunized via the intranasal (IN, n = 7) or tonsillar (IT, n = 7) route and control (n = 12) subjects were immunized via the intranasal route with E-GTF. Nasal wash, saliva, and serum were collected before immunization and then weekly for 3 months after immunization. Significant (P < 0.05) mucosal and serum immunoglobulin A (IgA) anti-E-GTF responses were observed in all three groups. Nasal and serum IgA anti-E-GTF responses were significantly higher (P < 0.05) in the IN group. The salivary responses in the three groups were, in general, similar. These results indicate that intranasal immunization primes the immune system for a localized secondary response to S. mutans antigens. © 2006 The Authors.