It is well known that the immune response in sheep against Brucella melitensis is subject to individual variation, depending on diverse factors. It bears asking whether these factors (e.g. clinical disease, active infection, state of previous immunity), when affecting a group, can cause variation in the performance of different diagnostic tests. To clarify some of the circumstances in which this immune response can vary, we examine the immune-response profile of sheep protected against the clinical disease by prior vaccination with strain Rev. 1 in comparison with the profile of unprotected females showing the classical brucellosis symptoms. An experimental infection was provoked at midpregnancy under controlled conditions of both non-vaccinated (n=7) and previously Rev.1-vaccinated ewes (n=5). Their immune response was monitored from 7 to 9 weeks before abortion or normal birth to 30 weeks afterwards. Antibody response was assessed by classical tests (Rose Bengal test, complement fixation test (CFT)) in comparison with other diagnostic tests (indirect ELISA (iELISA), competitive ELISA (cELISA), fluorescence polarization assay (FPA), immunocapture test (ICT)). In addition, the cell-mediated immune response was indirectly evaluated by the in vitro antigen-specific release of gamma-interferon. The antibody levels and antigen-specific gamma-IFN profile of the non-vaccinated ewes having the disease and excreting the pathogen was notably high and differed significantly (P<0.05 or P<0.01) from those of vaccinated ewes that neither contracted brucellosis nor excreted the pathogen. In general, all the tests detect the infection in the non-vaccinated ewes with substantial effectiveness. It can be concluded that the high levels of circulating antibodies and of antigen-specific gamma-IFN are related to active Brucella infection. Similarly, the state of protection against the disease, but not necessarily against infection, due to a previous immunization with the Rev. 1 vaccination, appears to be responsible for a low level of detectable immune response. Nevertheless, the design of the study limits conclusions to pregnant ewes and cannot be extrapolated to non-pregnant ewes or rams. Likewise, the study provides no information on animals which are carriers of B. melitensis.