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Using monoclonal antibodies in developing ELISA for diagnosing bovine tuberculosis.

Three stable strains of hybridoma cells (2H10, 3A8, 1B6) producing monoclonal antibodies (Mab) reacted with Protein Purified Derivative (PPD) Mycobacterium bovis were obtained. They were incapable of binding to PPD M. avium and whole cells M. bovis, M. kansasii, M. scrofulaceum as well as to their soluble and sonicated antigens. PPD M. bovis was attached to the solid phase via Mab which were directed to its protein with a molecular weight of 66 kDa. The diagnostic efficiency of sandwich ELISA was tested as compared with skin test. Blood samples were obtained from 849 cows belonging to farms unaffected and farms affected by tuberculosis. Ninety cows (10.6%) reacted to PPD M. bovis, but among them only 22 cows (2.6%) had a positive ELISA. Average titre of M. bovis specific antibodies detected in serum samples by ELISA was equal to 1:920 – 1:6400. Diagnostic value of ELISA and skin test was determined by autopsy of 12 cows. There were M. bovis specific antibodies at rather high titres in the blood samples of 9 cows which showed positive to skin test. Three cows developed a response to PPD M. bovis but in the blood samples of these animals we failed to detect any specific antibodies. Results of autopsy showed that all cows with positive ELISA and skin test had pathological findings in lungs and lymphatic nodes specific for tuberculosis, but internal organs of cows reacted only to skin test had no alterations. The results of our research testify to the high specificity and efficiency of Mab-based sandwich ELISA in isolating animals infected with tuberculosis and represent an evaluation of the actual epizootic situation.