STANDARDIZATION AND VALIDATION OF ELISA FOR DIAGNOSIS OF BOVINE LEPTOSPIROSIS

Leptospirosis is a zoonotic disease that causes serious health risks to animals. It is a disease of high economic impact, therefore the correct diagnosis and surveillance are essential to control infection. The aim of this study was to standardize and validate an indirect ELISA using outer membrane proteins (OMPs) of serovar Hardjo as the antigen of the test (ELISA-OMP/Hardjo), using the microscopic agglutination test (MAT) as a reference. The ELISA-OMP/Hardjo was standardized using 15 serum samples from positive animals and 15 negative in the MAT. The OMP / Hardjo was evaluated at concentrations of 0.35 mu g / mu L, 0.17 mu g / mu L, 0.08 mu g / mu L and 0.04 mu g / mu L. Serum samples were tested at dilutions 1:50, 1:100, 1:500, 1:1000, and anti IgG conjugate in 1:5000 and 1:10000. The validation of the test was performed using 218 serum samples. The standardization of ELISA-OMP/Hardjo indicated that the best protein concentration for sensitization of the plate was 0.08 mu g / mu L, 1:50 dilution of primary antibody and conjugated anti-IgG 1:5000. Of the 218 samples tested in MAT, 86 (39%) were positive and 132 (61%) negative. ELISA-OMP/Hardjo already identified 121 (55%) positive samples and 97 (45%) negative. The sensitivity was 100% and specificity 73%. Comparison of MAT and ELISA tests showed concordance of 0.83% and Kappa of 0.68 (p < 0.0001). The ELISA-OMP/Hardjo proved to be a sensitive test, indicating its potential as a screening tool in the diagnosis of bovine leptospirosis.