In this paper we report the development of a sensitive and specific assay for the detection of tuberculosis (TB) in European badgers (Meles meles), based on the stimulation of lymphocytes in whole-blood culture and the subsequent detection of gamma-interferon (IFNgamma) by sandwich ELISA. The comparative levels of IFNgamma produced to bovine and avian tuberculin (B-A) was used as the basis of determining the TB status of badgers, resulting in a more sensitive test than that based on the defined Mycobacterium bovis antigens ESAT6 and CFP10. The assay was evaluated using 235 badgers. The IFNgamma EIA (enzyme immunoassay) based on a monoclonal pair (mEIA) was more sensitive than one using a rabbit polyclonal antiserum (pEIA). At a specificity of 93.6%, the mEIA was 80.9% sensitive, compared to a sensitivity of 74.5% for the pEIA. At the same specificity as the EIA, the current serological ELISA test for TB in badgers (Brock test) had a sensitivity of 48.9%. Only one of the culture positive badgers missed by the mEIA was correctly diagnosed by the Brock test, suggesting that the combination of both a T-cell and serological test has little diagnostic advantage.