By immunizing mice with killed whole bacterial cells of Brucella Melitensis Rev1 a panel of four hybridomas producing monoclonal antibodies (mAb) specific for the surface antigens of this bacterium were produced. ELISA was used to screen the hybridoma supernatants. Immunoblots of the cell extract indicated that tow mAb were specific for S-LPS (BOX-1, BOX-2, and tow others were reactive with major outer membrane proteins (OMP) (Box-3, Box-4). The OMP recognized by these antibodies were the proteins with molecular masses of 22-25 kDa (Box-3, Box-4) and 30-33 kDa (Box-1). None of the three mAb including Box-2, Box-3 and Box-4 cross reacted with any other bacteria close to Brucella melitensis, but Box-1 cross reacted with B. abortus S19 and B. suis. By using cell extract and killed whole cell Ag in ELISA, it was indicated that all mAb except Box-1 have better reactivity with cell extract Ag, but Box-4 mAb reacted with killed whole cell Ag better than cell extract Ag. We utilize the isolated antibodies (Box4) in lateral flow test.