title: Development of a double-monoclonal antibody sandwich ELISA: Tool for chicken interferon-γ detection ex vivo.
abstract: The aim of the present work was to develop reagents to set up a chicken interferon-γ (ChIFN-γ) assay. Four monoclonal antibodies (mAbs) specific for ChIFN-γ were generated to establish sandwich ELISA based on 2 different mAbs. To improve the detection sensitivity of ChIFN-γ, a double-monoclonal antibody sandwich ELISA was developed using mAb 3E5 as capture antibody and biotinylated mAb 3E3 as a detection reagent. The results revealed that this ELISA has high sensitivity, allowing for the detection of 125 to 500 pg/mL of recombinant ChIFN-γ, and also has an excellent capacity for detecting native ChIFN-γ. This ELISA was then used to detect ChIFN-γ level in chickens immunized with a Newcastle disease virus (NDV) vaccine, the immunized chicken splenocytes were stimulated by NDV F protein as recall antigen. From our results, it appears that the sensitivity range of this sandwich ELISA test is adequate to measure the ex vivo release of ChIFN-γ.
authors.affiliation: Jiangsu Key Lab of Zoonosis, Yangzhou University, China (Dai, Xu, Chen, Chen X, Pan, Jiao); College of Medicine, Yangzhou University, China (Wang).
authors.forename: Hua
authors.initials: H
authors.lastname: Dai
authors.affiliation: Jiangsu Key Lab of Zoonosis, Yangzhou University, China (Dai, Xu, Chen, Chen X, Pan, Jiao); College of Medicine, Yangzhou University, China (Wang).
authors.forename: Zheng-Zhong
authors.initials: ZZ
authors.lastname: Xu
authors.affiliation: Jiangsu Key Lab of Zoonosis, Yangzhou University, China (Dai, Xu, Chen, Chen X, Pan, Jiao); College of Medicine, Yangzhou University, China (Wang).
authors.forename: Meiling
authors.initials: M
authors.lastname: Wang
authors.affiliation: Jiangsu Key Lab of Zoonosis, Yangzhou University, China (Dai, Xu, Chen, Chen X, Pan, Jiao); College of Medicine, Yangzhou University, China (Wang).
authors.forename: Jun-Hua
authors.initials: JH
authors.lastname: Chen
authors.affiliation: Jiangsu Key Lab of Zoonosis, Yangzhou University, China (Dai, Xu, Chen, Chen X, Pan, Jiao); College of Medicine, Yangzhou University, China (Wang).
authors.forename: Xiang
authors.initials: X
authors.lastname: Chen
authors.affiliation: Jiangsu Key Lab of Zoonosis, Yangzhou University, China (Dai, Xu, Chen, Chen X, Pan, Jiao); College of Medicine, Yangzhou University, China (Wang).
authors.forename: Zhi-Ming
authors.initials: ZM
authors.lastname: Pan
authors.affiliation: Jiangsu Key Lab of Zoonosis, Yangzhou University, China (Dai, Xu, Chen, Chen X, Pan, Jiao); College of Medicine, Yangzhou University, China (Wang).
authors.forename: Xin-An
authors.initials: XA
authors.lastname: Jiao
mesh_terms: D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology*; D002645:Chickens / Q000097:blood*; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods / Q000662:veterinary*; D005786:Gene Expression Regulation / Q000276:immunology; D007371:Interferon-gamma / Q000097:blood*; D051379:Mice; D008807:Mice, Inbred BALB C; D009521:Newcastle Disease / Q000517:prevention & control; D010957:Plasmids; D011994:Recombinant Proteins; D012680:Sensitivity and Specificity; D014765:Viral Vaccines / Q000276:immunology
chemical_list: D000911:Antibodies, Monoclonal; D011994:Recombinant Proteins; D014765:Viral Vaccines; D007371:Interferon-gamma