Recombinant OMP31Proteins Of Brucell Melitensis Strain Used As The Diagnostic Antigen In Indirect ELISA Test

In this study, recombinant omp31protein of Brucell melitensis strain was expressed for developing indirect ELISA for Brucell abortus strain differential diagnosis.The main results were as follows:1.Expression and immunogenicity of the recombinant omp31protein Brucell melitensis strainThe omp31gene were got from Brucella melitensis isolated in innermongolia by Polymerase chain reaction (PCR) and cloned into PMD-T. After sequencing, the omp31gene were cloned into vector pet-30a and expressed in Ecoli, then detected by SDS-PAGE and western-blot. The recombinant plasmid pet-30a-omp31was constructed successfully and expressed successfully in Ecoli. The recombinant protein could react with Brucella melitensis positive serum.2The Establishment of Indirect ELISA for detecting brucellosis infected by Brucell abortus strainThe indirect ELISA was developped by the recombinant0MP31antigen. The optimal result of ELISA as follow:coating concentration was2.5μg/ml(l/800). The optimal dilution of serum examined was1/100;1%BSA was determined to be the best blocking reagent of antigen,The optimal dillution of HRP were1:5000for ELISA; The cut-off OD450value of OMP31to judge result was0.337respectivelly. The recombinant OMP31protein showed no reactivity with antisera to B.abortus S19strain and negative serum. The result demonstrated that the coefficient of variability of OMP31base ELISA were10%and10%. Above result showed that the indirect ELISA method had advantages of good repetition, high sensibility, strong specificity, economic practicality and safety. So that, the indirect ELISA developed in this study might be widely used as a serological diagnostic method to brucellosis infected by Brucell abortus strain.