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| Preparation of Monoclonal Antibody Against σC Protein of Novel duck reovirus and Its Antigenic Epitope Identification |
| HUA Jiong-Gang, YE Wei-Cheng, NI Zheng, CHEN Liu, ZHU Yin-Chu, YUN Tao*, ZHANG Cun* |
| Institute of Animal Husbandry and Veterinary Sciences, Zhejiang Academy of Agriculture Sciences, Hangzhou 310021, China |
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Abstract Novel duck reovirus (NDRV) is a new pathogen discovered in recent years, which can cause diseases in multiple species of ducks and geese and huge economic losses to the duck industry in China. σC protein is one of the outer capsid proteins of NDRV, which is able to induce type-specific antibodies against the virus and functions in the identification and attachment of the virus to the target cell, probably via a receptor-mediated event. For the preparation of monoclonal antibody (McAb) against protein σC of NDRV and identification of antigen epitope, SP2/0 myeloma cells were fused with spleen from BALB/c mice (Mus musculus) immunized with purified recombinant σC expressed in Escherichia coli. Four hybidoma cell lines (A5-A1, A5-B6, A9-D4 and B9-6) stably secreting McAbs against σC of NDRV were identified by indirect ELISA. Western blot results showed that all 4 McAbs could recognize whole virus of NDRV and recombinant σ C protein from prokaryotic expression. Indirect immunofluorescence assay (IFA) indicated that 4 McAbs reacted positively with NDRV, but did not show cross reaction with Classical duck reovirus (CDRV) and Avian reovirus (ARV). The antigenic epitopes recognized by 4 McAbs were identified by synthetic overlapping peptide library and a series of truncated peptides, and the results showed that the core antigenic epitope of 177PILSGPADA185 was recognized by 3 McAbs (A5-A1, A5-B6, and B9-6). The antigenic epitopes recognized by A9-D4 might be spatial conformational epitopes. Conservative analysis showed that the antigenic epitope was highly conservative in the NDRV isolates from different regions and different hosts. The results suggested that 177PILSGPADA185 was a type-specific linear B-cell epitope of NDRV. This study could provide a basis for further research on the structure of σC protein and the establishment of NDRV clinical detection methods.
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Received: 23 April 2021
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Corresponding Authors:
*yt-t@163.com; zhangcun@aliyun.com
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