Indirect ELISA for diagnosis of Brucella ovis infection in rams

Brucella ovis is a major cause of epididymitis in sexually mature rams, resulting in subfertility, infertility, and economic losses for the sheep industry worldwide. The aim of this study was to develop an indirect ELISA (iELISA) using recombinant proteins, namely rBoP59 and rBP26, as antigens for serological diagnosis of B. ovis infection. The BoP59 and BP26 recombinant proteins were expressed in E. coli and purified by affinity chromatography. Antigenicity was tested by Western blot and iELISA. Standardization of iELISA was performed with 500ng and 1µg BoP59 and rBP26 per well, testing serum from uninfected and experimentally infected rams. rBP26 was effective in distinguishing positive from negative rams. The rBP26 iELISA developed in this study is the first to use a completely purified rBP26 as antigen resulting in high sensitivity (100%) and specificity (90.2%), and an overall accuracy equal to 1.0.