Characterization of Monoclonal Antibodies against σA Protein and Cross-Reactive Epitope Identification and Application for Detection of Duck and Chicken Reovirus Infections
Although σA is an important major core protein of duck reovirus (DRV), the B-cell epitopes of this protein remain unknown to reseacrhers. Six monoclonal antibodies (MAbs) (1A7, 3F4, 5D2, 4E2, 3C7, and 2B7) were developed by using prokaryotic-expressed recombinant His-σA protein. Fi...
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MDPI AG
2019-09-01
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| Series: | Pathogens |
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| Online Access: | https://www.mdpi.com/2076-0817/8/3/140 |
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| author | Xueming Chen Tongtong Li Xiaodan Chen Chenxi Li Weiwei Lin Hongyu Liu Shuping Song Xiaofei Bai Yun Zhang |
| author_facet | Xueming Chen Tongtong Li Xiaodan Chen Chenxi Li Weiwei Lin Hongyu Liu Shuping Song Xiaofei Bai Yun Zhang |
| author_sort | Xueming Chen |
| collection | DOAJ |
| description | Although σA is an important major core protein of duck reovirus (DRV), the B-cell epitopes of this protein remain unknown to reseacrhers. Six monoclonal antibodies (MAbs) (1A7, 3F4, 5D2, 4E2, 3C7, and 2B7) were developed by using prokaryotic-expressed recombinant His-σA protein. Five of six MAbs (1A7, 3F4, 4E2, 3C7, and 2B7) reacted with His-σA protein in a conformation-independent manner, while 5D2 reacted with σA in a conformation-dependent manner. Immunofluorescence assays showed that the MAbs could specifically bind to DRV infected BHK-21 cells. The MAbs were delineated as three groups by a competitive binding assay. By using 12-mer peptide phage display and mutagenesis, MAb 4E2 was identified to recognize minimal epitope <sup>56</sup>EAPYPG<sup>61</sup> and MAb 1A7 recognize <sup>341</sup>WVV/MAGLI/V<sup>347</sup>, residues <sup>341</sup>V/M and <sup>347</sup>I/V are replaceable. Dot blotting and sequence analysis confirmed that EAPYPG and WVV/MAGLI/V are cross-reactive epitopes in both DRV and avian reovirus (ARV). An enzyme-linked immunosorbent assay (ELISA) based on two expressed EAPYPG and WVVAGLI as antigen demonstrated its diagnostic potential by specific reacting with serum samples from DRV- or ARV-infected birds. Based on these observations, an epitope-based ELISA could be potentially used for DRV or ARV surveillance. These findings provide insights into the organization of epitopes on σA protein that might be valuable for the development of epitope-based serological diagnostic tests for DRV and ARV infection. |
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| language | English |
| last_indexed | 2024-04-11T13:57:09Z |
| publishDate | 2019-09-01 |
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| spelling | doaj.art-7aba56eb1546490b86a2ee9a913429e52022-12-22T04:20:13ZengMDPI AGPathogens2076-08172019-09-018314010.3390/pathogens8030140pathogens8030140Characterization of Monoclonal Antibodies against σA Protein and Cross-Reactive Epitope Identification and Application for Detection of Duck and Chicken Reovirus InfectionsXueming Chen0Tongtong Li1Xiaodan Chen2Chenxi Li3Weiwei Lin4Hongyu Liu5Shuping Song6Xiaofei Bai7Yun Zhang8State Key Laboratory of Veterinary Biotechnology, Harbin Veterinary Research Institute of Chinese Academy of Agricultural Sciences, Harbin 150069, ChinaState Key Laboratory of Veterinary Biotechnology, Harbin Veterinary Research Institute of Chinese Academy of Agricultural Sciences, Harbin 150069, ChinaState Key Laboratory of Veterinary Biotechnology, Harbin Veterinary Research Institute of Chinese Academy of Agricultural Sciences, Harbin 150069, ChinaState Key Laboratory of Veterinary Biotechnology, Harbin Veterinary Research Institute of Chinese Academy of Agricultural Sciences, Harbin 150069, ChinaState Key Laboratory of Veterinary Biotechnology, Harbin Veterinary Research Institute of Chinese Academy of Agricultural Sciences, Harbin 150069, ChinaState Key Laboratory of Veterinary Biotechnology, Harbin Veterinary Research Institute of Chinese Academy of Agricultural Sciences, Harbin 150069, ChinaState Key Laboratory of Veterinary Biotechnology, Harbin Veterinary Research Institute of Chinese Academy of Agricultural Sciences, Harbin 150069, ChinaState Key Laboratory of Veterinary Biotechnology, Harbin Veterinary Research Institute of Chinese Academy of Agricultural Sciences, Harbin 150069, ChinaState Key Laboratory of Veterinary Biotechnology, Harbin Veterinary Research Institute of Chinese Academy of Agricultural Sciences, Harbin 150069, ChinaAlthough σA is an important major core protein of duck reovirus (DRV), the B-cell epitopes of this protein remain unknown to reseacrhers. Six monoclonal antibodies (MAbs) (1A7, 3F4, 5D2, 4E2, 3C7, and 2B7) were developed by using prokaryotic-expressed recombinant His-σA protein. Five of six MAbs (1A7, 3F4, 4E2, 3C7, and 2B7) reacted with His-σA protein in a conformation-independent manner, while 5D2 reacted with σA in a conformation-dependent manner. Immunofluorescence assays showed that the MAbs could specifically bind to DRV infected BHK-21 cells. The MAbs were delineated as three groups by a competitive binding assay. By using 12-mer peptide phage display and mutagenesis, MAb 4E2 was identified to recognize minimal epitope <sup>56</sup>EAPYPG<sup>61</sup> and MAb 1A7 recognize <sup>341</sup>WVV/MAGLI/V<sup>347</sup>, residues <sup>341</sup>V/M and <sup>347</sup>I/V are replaceable. Dot blotting and sequence analysis confirmed that EAPYPG and WVV/MAGLI/V are cross-reactive epitopes in both DRV and avian reovirus (ARV). An enzyme-linked immunosorbent assay (ELISA) based on two expressed EAPYPG and WVVAGLI as antigen demonstrated its diagnostic potential by specific reacting with serum samples from DRV- or ARV-infected birds. Based on these observations, an epitope-based ELISA could be potentially used for DRV or ARV surveillance. These findings provide insights into the organization of epitopes on σA protein that might be valuable for the development of epitope-based serological diagnostic tests for DRV and ARV infection.https://www.mdpi.com/2076-0817/8/3/140duck reovirusavian reovirusturkey reovirusmonoclonal antibodyσA protein epitopescross-reactive epitopesdiagnosis |
| spellingShingle | Xueming Chen Tongtong Li Xiaodan Chen Chenxi Li Weiwei Lin Hongyu Liu Shuping Song Xiaofei Bai Yun Zhang Characterization of Monoclonal Antibodies against σA Protein and Cross-Reactive Epitope Identification and Application for Detection of Duck and Chicken Reovirus Infections Pathogens duck reovirus avian reovirus turkey reovirus monoclonal antibody σA protein epitopes cross-reactive epitopes diagnosis |
| title | Characterization of Monoclonal Antibodies against σA Protein and Cross-Reactive Epitope Identification and Application for Detection of Duck and Chicken Reovirus Infections |
| title_full | Characterization of Monoclonal Antibodies against σA Protein and Cross-Reactive Epitope Identification and Application for Detection of Duck and Chicken Reovirus Infections |
| title_fullStr | Characterization of Monoclonal Antibodies against σA Protein and Cross-Reactive Epitope Identification and Application for Detection of Duck and Chicken Reovirus Infections |
| title_full_unstemmed | Characterization of Monoclonal Antibodies against σA Protein and Cross-Reactive Epitope Identification and Application for Detection of Duck and Chicken Reovirus Infections |
| title_short | Characterization of Monoclonal Antibodies against σA Protein and Cross-Reactive Epitope Identification and Application for Detection of Duck and Chicken Reovirus Infections |
| title_sort | characterization of monoclonal antibodies against σa protein and cross reactive epitope identification and application for detection of duck and chicken reovirus infections |
| topic | duck reovirus avian reovirus turkey reovirus monoclonal antibody σA protein epitopes cross-reactive epitopes diagnosis |
| url | https://www.mdpi.com/2076-0817/8/3/140 |
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