The immunological diagnosis of Johne's disease is performed by an ELISA test and Johnin test. For the production of the diagnostic antigen for the tests, P-18 strain, classified as Mycobacterium avium subsp. avium(Maa) on the basis of genetic properties, has been used. Hence, in the present study, we used Kag-1 strain of Mycobacterium avium subsp. Paratuberculosis(Map), an isolate from Japan, to produce an ELISA antigen and examined its sensitivity and specificity in antibody detection.

Sera from 141 fecal culture-positive and 103 fecal culture-negative cattle were used to examine the sensitivity and precision of the partially purified ELISA antigen from Kag-1 strain of Map. The sensitivity was 80.1% in ELISA and an agreement of 88.1% was found between the ELISA method and fecal culture method (Kappa value: 0.765, P<0.01). In addition, in the fecal culture-positive cattle, the ELISA-positive ratio was 71.8% for light shedders, 80.9% for intermediate shedders, and 92.9% for heavy shedders. The average ratio was 79.4%. In examination of cross reactions with bovine Mycobacterium antibodies, the purified ELISA antigen revealed week cross reactions with antisera immunized by BCG Tokyo strain of M. bovis and Kumamoto-8 strain of Maa, and a strong cross reaction with an antiserum immunized by S-7 strain of M. intracellulare. A western blotting that uses infection serum of ATCC 19698 strain and Kag-1 strain of Map showed colored bands mainly at 56 kDa and 29 kDa.

The above results indicate that the purified protein from Kag-1 strain of Map can be used as an antigen for ELISA.