Genetic fusion of tandem repeats of the complement molecule C3d has been shown to considerably enhance immune responses to genetic vaccines. We have investigated the applicability of this approach to augment humoral immune responses toward vaccines delivered by recombinant adeno-associated virus (AAV) vectors. C3d(3)-fusion was found to markedly decrease antibody responses to merozoite surface protein 4/5 from Plasmodium yoelii and contrasted with greater than 50-fold enhancement in responses when this strategy was similarly applied to another AAV-encoded model antigen, hen egg lysozyme. These data indicate that the efficacy of the C3d(3) strategy operates in an antigen-dependent manner. Additional studies also showed that homologous recombination events between the C3d tandem repeats occurred during vector packaging and transduction resulting in expression of C3d(1)-, C3d(2)-, C3d(3)- and C3d(4)-fused antigen. This is the first report to apply the C3d approach to augment responses against a recombinant viral vector system and the consequences of these findings are discussed.