Expression And Purification Of Brucella Outer Membrane Protein2a And Establish Stable Hybridoma Cell Lines Secreting Specific Antibody

Brucellosis is a chronic zoonosis caused by Brucella, which is a genus of gram-negative,intracellular facultative bacteria. The disease mainly caused infertility, miscarriage andstillbirth. It resulted in the economic loss of breeding industry and the threatened to humanlife. Over the years, scholars have studied outer membrane proteins of Brucella and found thatthe major outer membrane protein is immunogenic and protective antigen. Omp2a belongs tothe second group of outer membrane protein of Brucella, is one kind of channel protein in thecell surface. The second set outer membrane proteins constitute the penetration system ofBrucella that can cause fluid and cellular immune responses. So, for the sake of exploring anew method of diagnosis, which is faster and more accurate for detection of brucellosis, wedid this study to obtain the stable hybridoma cell line that can product the antibody of Omp2aby expressed and purified Omp2a, immunized mice, cell fusion and screening.1. We used the prokaryotic expression vector with GST tag to express the Brucella outermembrane protein2a, and purified by electrical elution.2. The purified Omp2a fusion protein was used to immunize Balb/c mice. Indirect ELISAmethod for detection of immune serum antibody titer in mice, we should proceed cell fusionwhile the antibody titer reached1∶1×105.3. We obtained three stable hybridoma cell line which secret specific antibody by cell fusionand screening.The result of this experiment can provide certain material foundation and technicalsupport for the further study of new vaccine and diagnostic.