A monoclonal antibody directed against the largest bluetongue virus (BTV) non-structural protein (NS1) was used in a competitive ELISA to detect antibodies to tubules (composed of NS1) in serum samples. Anti-tubule antibodies were detected at approximately 10 days post-infection in BTV infected sheep but no such antibodies were detected in sheep injected with inactivated BTV vaccines. Tubules are also produced during the replication of other related orbiviruses, including epizootic haemorrhagic disease of deer virus (EHDV). However, antibodies to the EHDV tubules were not detected in antisera from EHDV infected animals using the BTV NS1 specific monoclonal antibody and this assay system, confirming the serogroup-specific nature of this assay. This assay may prove useful not only for confirming the inactivated nature of experimental vaccines but also for the detection of viral replication and hence measurement of protection in vaccine potency trials following virulent BTV challenge. If inactivated BTV vaccines are ever widely adopted this test could provide a valuable means of discriminating between vaccinated and infected animals.