title: Application of recombinant bovine viral diarrhea virus proteins in the diagnosis of bovine viral diarrhea infection in cattle.
abstract: The National Animal Disease Laboratory (NADL) vaccine strain of bovine viral diarrhea virus (BVDV) genes for gp48 and p80 were expressed in Escherichia coli. The BVDV-NADL gene for gp62 was integrated into a baculovirus genome for expression in Spodoptera frugiperda (Sf-9) insect ovarian cells. The antigenicity of baculovirus expressed BVDV protein was detected by anti-BVDV specific antibodies in an enzyme-linked immunosorbent assay (ELISA), indirect immunofluorescent assay (IFA) and radio-immunoprecipitation (RIP). The recombinant proteins isolated from bacteria showed antigenic properties when analyzed by ELISA and immunoblotting using BVDV antibodies. The recombinant proteins were then used in ELISA or IFA to detect BVDV infection by testing 54 independent bovine serum samples. The baculovirus-expressed BVDV protein was used as an ELISA and IFA antigen, and the bacteria-expressed proteins were used as ELISA antigens. BVDV-NADL-infected Madin-Darby bovine kidney (MDBK) cell monolayers served as a control antigen. Statistical analysis showed a high degree of correlation between the reactivity of recombinants and natural antigens in ELISA using bovine sera. The results of ELISA or IFA proved there is a high degree of correlation with the virus neutralization. In the comparative ELISA assays, the insect-cell-mediated expression revealed greater specificity and sensitivity than the bacterial expression or the natural BVDV antigens produced by cell cultures.
authors.affiliation: Department of Diagnostic Medicine/Pathobiology, College of Veterinary Medicine, Manhattan, KS 66506, USA.
authors.forename: J R
authors.initials: JR
authors.lastname: Reddy
authors.forename: J
authors.initials: J
authors.lastname: Kwang
authors.forename: O
authors.initials: O
authors.lastname: Okwumabua
authors.forename: S
authors.initials: S
authors.lastname: Kapil
authors.forename: T M
authors.initials: TM
authors.lastname: Loughin
authors.forename: K F
authors.initials: KF
authors.lastname: Lechtenberg
authors.forename: M M
authors.initials: MM
authors.lastname: Chengappa
authors.forename: H C
authors.initials: HC
authors.lastname: Minocha
mesh_terms: D000818:Animals; D000911:Antibodies, Monoclonal; D000914:Antibodies, Viral / Q000097:blood*; D000956:Antigens, Viral / Q000096:biosynthesis*; D001912:Bovine Virus Diarrhea-Mucosal Disease / Q000175:diagnosis* / Q000276:immunology; D002417:Cattle; D003001:Cloning, Molecular; D004797:Enzyme-Linked Immunosorbent Assay; D004926:Escherichia coli; D019084:Fluorescent Antibody Technique, Indirect; D005814:Genes, Viral; D009500:Neutralization Tests; D010576:Pestivirus / Q000235:genetics*; D011863:Radioimmunoassay; D011994:Recombinant Proteins / Q000096:biosynthesis; D018411:Spodoptera; D014162:Transfection; D014764:Viral Proteins / Q000096:biosynthesis*
chemical_list: D000911:Antibodies, Monoclonal; D000914:Antibodies, Viral; D000956:Antigens, Viral; D011994:Recombinant Proteins; D014764:Viral Proteins