In this study, novel serological tests were used to detect tuberculosis (TB) in groups of farmed red deer (Cervus elaphus) varying in disease status or possible confounding factors. Groups of deer naturally or experimentally infected with Mycobacterium bovis and animals vaccinated against paratuberculosis were studied, as were uninfected animals and animals naturally or experimentally infected with Mycobacterium avium subsp. paratuberculosis. Sera were assayed using two rapid lateral-flow tests, Chembio's CervidTB STAT-PAK and DPP VetTB tests, and results were compared to those from tuberculin skin tests. Both serological tests had a high sensitivity, but specificity was adversely affected after animals had received a vaccine against paratuberculosis and were subsequently skin tested. The specificity of the DPP VetTB test was higher than that of the CervidTB STAT-PAK test, with natural infection with M. avium subsp. paratuberculosis adversely affecting the specificity of only the CervidTB STAT-PAK test. The sera from M. avium subsp. paratuberculosis-infected deer that produced false-positive reactions in the CervidTB STAT-PAK test were retested with a multiantigen print immunoassay (MAPIA), and some of these sera were shown to react with the MPB83 antigen. Combining the results from the serological tests and the skin tests showed only a slight increase in the sensitivity of detection of M. bovis-infected animals. It is concluded that both the CervidTB STAT-PAK and DPP VetTB tests offer rapid, convenient, and easy detection of bovine tuberculosis in deer, albeit with significant interference from paratuberculosis vaccination status and subsequent skin testing. The latter finding illustrates one of the limitations of currently available vaccines against paratuberculosis.Infections with Mycobacterium bovis in farmed or wild deer continue to be a problem in many areas of the world, and novel, more efficient tests aimed at diagnosis of this disease are needed (1, 5). One of the main tools used as a diagnostic approach is the skin test procedure using purified protein derivative (PPD) from M. bovis (1). Unfortunately, the tuberculin skin test in deer has a relatively low specificity, and it is considered prudent to retest skin test-positive animals by using a confirmatory test to avoid the slaughter of false-positive responders (2, 11). It has been suggested that the high prevalence of paratuberculosis in farmed deer causes the cross-reactivity in the skin test, leading to a high rate of false positives (2, 11). Alternative detection strategies have been explored, but these alternatives are costly or time-consuming (15,16,17). An IgG1 enzyme-linked immunosorbent assay (ELISA) for detection of tuberculosis (TB) in deer has been developed, but results were confounded by infection with Mycobacterium avium subsp. paratuberculosis, the causative agent of paratuberculosis (12). Vaccination against paratuberculosis also confounds tests for TB in deer (13), and it is important to assess whether the use of any new serological ...