In this study we firstly established a vaccination/challenge model to study pseudorabies virus infection in mice. The mouse model was used to investigate the significance of CD4 and CD8 cells and of IFN production in protective immunity. Functional depletion of CD4 and CD8 and IFN was obtained in vivo by intraperitoneal injection of alginate-encapsulated anti-CD4, -CD8 or -IFN producing hybridoma's before and at the moment of vaccination. The observed protective immunity was correlated with underlying immunologic responses such as PRV-specific DTH reactivity, lymphoproliferation and cytotoxicity. The significance of CD4 and CD8 cells and of IFN production was also investigated for these immunological responses by the same in vivo depletion technique. The results demonstrated that protective vaccination of mice, that could be induced by immunization with 107 plaque forming units of the avirulent PRV mutant NIA3 TK−, was characterized by a typical anti-viral Th1 type immune response. A clear PRV-specific, CD4-dependent DTH reactivity and a classical CD8-dependent, MHC-restricted cytotoxicity was induced after protective immunization and the humoral immune response had a bias towards PRV-specific IgG2a formation. In vivo treatment with anti-CD8 and anti-IFN demonstrated that the cytotoxic response and humoral IgG2a response, respectively, were strongly reduced, whereas protection against lethal challenge was unaffected. On the other hand anti-CD4 treatment reduced the induced protection so that 30 f the mice died after lethal challenge. The results of our study demonstrated that CD4 , DTH like effector cells are a crucial effector mechanism for protective immunity against PRV.