title: Generation of E. coli-derived virus-like particles of porcine circovirus type 2 and their use in an indirect IgG enzyme-linked immunosorbent assay.
abstract: Porcine circovirus type 2 (PCV2) causes increased mortality and poor growth or weight loss in apparently healthy swine. Therefore, methods to detect PCV2-specific antibodies in swine serum are important for prevention, diagnosis, and control of PCV2-associated diseases (PCVAD). In this study, PCV2 virus-like particles (VLPs) were used to develop a rapid, simple and economical indirect enzyme-linked immunosorbent assay to detect (with high sensitivity) PCV2-specific antibodies in swine serum. The PCV2 capsid protein (Cap) was overexpressed in E. coli after optimizing the cap gene. Subsequently, the soluble Cap was rapidly purified in one step by automated fast protein liquid chromatography (FPLC). The purified PCV2 Cap was shown by transmission electron microscopy and gel filtration chromatography to be capable of self-assembling into VLPs in vitro. Using the purified VLPs as antigens, optimal operating conditions for the VLP ELISA were determined. The concentration of PCV2 VLPs was 1 µg/ml per well, and the dilution factors for swine serum and horseradish peroxidase (HRP)-labeled goat anti-pig antibody were 1:150 and 1:4000, respectively. Out of 241 serum samples tested with this assay, 83.4 % were found to be positive. Importantly, the VLP ELISA had a total coincidence rate of 97.4 % (74/76) compared to an Ingezim PCV2 ELISA IgG assay. In summary, this rapid, inexpensive VLP ELISA has the potential to greatly facilitate large-scale investigations of PCV2-associated serotypes.
authors.affiliation: Laboratory of Functional Proteomics (LFP) and Research Center of Reverse Vaccinology (RCRV), College of Veterinary Medicine, Hunan Agricultural University, Furong District, Changsha, 410128, People's Republic of China.
authors.forename: Yan
authors.initials: Y
authors.lastname: Zhang
authors.affiliation: Laboratory of Functional Proteomics (LFP) and Research Center of Reverse Vaccinology (RCRV), College of Veterinary Medicine, Hunan Agricultural University, Furong District, Changsha, 410128, People's Republic of China.
authors.forename: Zhanfeng
authors.initials: Z
authors.lastname: Wang
authors.affiliation: Laboratory of Functional Proteomics (LFP) and Research Center of Reverse Vaccinology (RCRV), College of Veterinary Medicine, Hunan Agricultural University, Furong District, Changsha, 410128, People's Republic of China.
authors.forename: Yang
authors.initials: Y
authors.lastname: Zhan
authors.affiliation: Laboratory of Functional Proteomics (LFP) and Research Center of Reverse Vaccinology (RCRV), College of Veterinary Medicine, Hunan Agricultural University, Furong District, Changsha, 410128, People's Republic of China.
authors.forename: Qian
authors.initials: Q
authors.lastname: Gong
authors.affiliation: Laboratory of Functional Proteomics (LFP) and Research Center of Reverse Vaccinology (RCRV), College of Veterinary Medicine, Hunan Agricultural University, Furong District, Changsha, 410128, People's Republic of China.
authors.forename: Wanting
authors.initials: W
authors.lastname: Yu
authors.affiliation: Laboratory of Functional Proteomics (LFP) and Research Center of Reverse Vaccinology (RCRV), College of Veterinary Medicine, Hunan Agricultural University, Furong District, Changsha, 410128, People's Republic of China.
authors.forename: Zhibang
authors.initials: Z
authors.lastname: Deng
authors.affiliation: Laboratory of Functional Proteomics (LFP) and Research Center of Reverse Vaccinology (RCRV), College of Veterinary Medicine, Hunan Agricultural University, Furong District, Changsha, 410128, People's Republic of China.
authors.forename: Aibing
authors.initials: A
authors.lastname: Wang
authors.affiliation: Laboratory of Functional Proteomics (LFP) and Research Center of Reverse Vaccinology (RCRV), College of Veterinary Medicine, Hunan Agricultural University, Furong District, Changsha, 410128, People's Republic of China.
authors.forename: Yi
authors.initials: Y
authors.lastname: Yang
authors.affiliation: Laboratory of Functional Proteomics (LFP) and Research Center of Reverse Vaccinology (RCRV), College of Veterinary Medicine, Hunan Agricultural University, Furong District, Changsha, 410128, People's Republic of China. naidongwang@hunau.edu.cn.
authors.forename: Naidong
authors.initials: N
authors.lastname: Wang
mesh_terms: D000818:Animals; D000914:Antibodies, Viral / Q000097:blood; D036022:Capsid Proteins / Q000235:genetics / Q000276:immunology; D018173:Circoviridae Infections / Q000276:immunology / Q000662:veterinary; D017925:Circovirus / Q000145:classification / Q000235:genetics / Q000276:immunology*; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods / Q000662:veterinary*; D004926:Escherichia coli / Q000235:genetics / Q000276:immunology; D007074:Immunoglobulin G / Q000097:blood; D065288:Serogroup; D034421:Sus scrofa; D013552:Swine / Q000276:immunology; D013553:Swine Diseases / Q000276:immunology / Q000821:virology; D058425:Vaccines, Virus-Like Particle / Q000235:genetics / Q000276:immunology
chemical_list: D000914:Antibodies, Viral; D036022:Capsid Proteins; D007074:Immunoglobulin G; D058425:Vaccines, Virus-Like Particle