Expression And Purification Of Recombinant Omp2 Of Chlamydia Trachomatis And Preparation And Identification Of Monoclonal Antibody

Objective:To express and purify outer membrane protein 2(Omp2) 167~434 amino acid residues of Chlamydia trachomatis serovar D in E. coli BL21(DE3), and to prepare and identify Monoclonal Antibodies Against recombinant 0mp2 of Chlamydia trachomatis.Methods:Recombinant 0mp2 (rOmp2') was expressed after induced by IPTG and analyzed by SDS-PAGE and Western blot, and purified with Ni-NTA-His affinity chromatography. The purification protein was refolded in GSH-GSSH buffer by stepwise gradient reducing the concentration of denaturant.Protein concentration was determined by Bradford assay. Hybridoma lines were obtained by fusing myeloma SP2/0 and spleen cells from BALB/c mouse immuned with CT rOmp2'.By means of indirect ELISA selection and several subcloning with limited dilution method,a monoclonal antibody (MAb) was produced. The characters of hybiridoma were dectected by FCM test and counting chromosomes.That was shown to be subclass by the double immunodiffusion analysis. The antibody was proved to be specific to CT Omp2 by western blot.Results:The rOmp2'was observed as a band of molecular relative weight of 35KDa or so by a SDS-PAGE gel and mainly existed in the pattern of inclusion body. After...