title: Real-time SPR characterization of the interactions between multi-epitope proteins and antibodies against classical swine fever virus.
abstract: The envelope glycoprotein E2 is the major immunodominant protein of the classical swine fever virus and can induce neutralizing antibodies and protective host-immune responses in infected swine. We designed, expressed, and purified multi-epitope protein (GST-BT22) that contains a tandem repeat of the E2 antigenic-determinant residues 693-704, 770-780, and 826-843, each of which is separated by a GGSSGG sequence. In the same manner, we also designed, expressed, and purified a second protein (GST-BT23) that contains a C-terminal sequence consisting of residues 1446-1460 from the classical swine fever virus nonstructural protein NS2-3 separated from the GST-BT22 sequence by a GGSSGG sequence. Western blotting of GST-BT22 and GST-BT23 with serum from a swine that had been experimentally infected with the virus showed that the proteins reacted with anti-serum, whereas GST did not. Surface plasmon resonance was used to quantify the affinities of GST-BT22 and GST-BT23 for serum antibodies (K(a) = 4.31 × 10(8) and 5.01 × 10(8), respectively). GST, used as a control, was reacted an order of magnitude less strongly than did GST-BT22 and GST-BT23. Surface plasmon resonance, therefore, appears to be a sensitive and precise method for epitope evaluation and can be used to characterize the immunogenicity of a recombinant protein.
authors.affiliation: Key Laboratory of Animal Virology of the Ministry of Agriculture, State Key Laboratory of Veterinary Etiological Biology, Lanzhou Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Lanzhou, Gansu 730046, China. ltth1977@163.com
authors.forename: Hong
authors.initials: H
authors.lastname: Tian
authors.forename: Xiangming
authors.initials: X
authors.lastname: Hou
authors.forename: Xiangtao
authors.initials: X
authors.lastname: Liu
mesh_terms: D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology; D057134:Antibodies, Neutralizing / Q000276:immunology*; D000914:Antibodies, Viral / Q000276:immunology*; D000937:Antigen-Antibody Reactions; D006691:Classical Swine Fever / Q000097:blood / Q000276:immunology; D006692:Classical Swine Fever Virus / Q000276:immunology*; D016056:Immunodominant Epitopes / Q000276:immunology*; D020349:Surface Plasmon Resonance / Q000379:methods*; D013552:Swine / Q000276:immunology / Q000821:virology*; D013997:Time Factors; D014759:Viral Envelope Proteins / Q000276:immunology*; D014765:Viral Vaccines / Q000276:immunology
chemical_list: D000911:Antibodies, Monoclonal; D057134:Antibodies, Neutralizing; D000914:Antibodies, Viral; D016056:Immunodominant Epitopes; D014759:Viral Envelope Proteins; D014765:Viral Vaccines; C087086:glycoprotein E2, classical swine fever virus