title: Expression, purification, and improved antigenic specificity of a truncated recombinant bp26 protein of Brucella melitensis M5-90: a potential antigen for differential serodiagnosis of brucellosis in sheep and goats.
abstract: Antibodies produced in animals vaccinated using live attenuated vaccines against Brucella spp. are indistinguishable using current conventional serological tests from those produced in infected animals. One potential approach is to develop marker vaccines in which specific genes have been deleted from parental vaccine strains that show good immunogenicity and vaccine efficacy. Corresponding methods of detection for antibodies raised by the marker vaccine should also be developed. A specific fragment of the bp26 gene of Brucella melitensis M5-90 was cloned into vector pQE32 to construct the recombinant plasmid (pQE32-rΔbp26). It was used to transform Escherichia coli M15 (pREP4) host cells, which expressed the rΔbp26 protein. Subsequently, the recombinant protein was purified by immobilized metal affinity chromatography and size-exclusion chromatography. The results of sodium dodecyl sulfate-polyacrylamide gel electrophoresis showed that the purified rΔbp26 protein was represented by only one band, with a molecular weight of 14 kDa, and it showed good antigenic specificity on western blot and enzyme-linked immunosorbent assay (ELISA). The purified rΔbp26 protein was intended to be used as an antigen to develop a novel ELISA to differentiate animals vaccinated with bp26 mutants of Brucella spp. from those infected naturally and those vaccinated with the parental vaccine strains.
authors: College of Veterinary Medicine, Nanjing Agricultural University, Nanjing, P R China.
authors: Wen-xing
authors: WX
authors: Liu
authors: Sen
authors: S
authors: Hu
authors: Zu-jian
authors: ZJ
authors: Qiao
authors: Wei-ye
authors: WY
authors: Chen
authors: Lin-tao
authors: LT
authors: Liu
authors: Fang-kun
authors: FK
authors: Wang
authors: Rong-hong
authors: RH
authors: Hua
authors: Zhi-gao
authors: ZG
authors: Bu
authors: Xiang-rui
authors: XR
authors: Li
mesh_terms: D000818:Animals; D002004:Brucella Vaccine / Q000235:genetics; D017347:Brucella melitensis / Q000235:genetics / Q000378:metabolism*; D002006:Brucellosis / Q000175:diagnosis / Q000276:immunology / Q000662:veterinary; D004797:Enzyme-Linked Immunosorbent Assay; D000939:Epitopes / Q000276:immunology; D015964:Gene Expression Regulation, Bacterial; D015511:Goat Diseases / Q000097:blood / Q000175:diagnosis / Q000276:immunology; D006041:Goats; D008565:Membrane Proteins / Q000276:immunology / Q000302:isolation & purification; D011994:Recombinant Proteins / Q000276:immunology / Q000302:isolation & purification; D012698:Serologic Tests / Q000379:methods*; D012756:Sheep; D012757:Sheep Diseases / Q000175:diagnosis / Q000276:immunology
chemical_list: C106040:BP26 protein, Brucella; D002004:Brucella Vaccine; D000939:Epitopes; D008565:Membrane Proteins; D011994:Recombinant Proteins