Mycobacterium paratuberculosis causes Johne's disease, an economically significant problem in ruminants in most countries. Serological tests for M. ptb. are useful in estimating the prevalence of infection when it is established in herds/flocks and for the confirmation of diagnosis in animals showing clinical signs. The present study was designed to evaluate a commercial ELISA kit (Institut Pourquier) under Australian conditions and to compare it with the existing agar gel immunodiffusion (AGID) test for the diagnosis of ovine and caprine paratuberculosis. Serum samples from 2200 uninfected sheep and 946 uninfected goats varying in breed, age and geographic locations were tested to estimate diagnostic specificity for each species. The diagnostic sensitivity of assays for sheep was estimated by testing 2550 sheep serum samples from various infected properties, while for goats, 125 serum samples from infected animals were tested. Results for plates were accepted or all samples on the plate were retested according to decision-limit criteria for positive and negative control sera, while individual samples with a coefficient of variation (CV) for paired replicates of more than 15% were also retested. The CV of positive and negative control sera was 9.7 and 11.4%, respectively. Diagnostic specificity and sensitivity were calculated at different cut point ratios and results were analysed by age, geographic location, and histological lesion type and culture results