Pigs are susceptible to foot-and-mouth disease virus (FMDV), and the humoral immune response plays an essential role in protection against FMDV infection. However, little information is available about FMDV-specific mAbs derived from single B cells of pigs. This study aimed to determine the antigenic features of FMDV that are recognized by antibodies from pigs. Therefore, a panel of pig-derived mAbs against FMDV were developed using fluorescence-based single B cell antibody technology. Western blotting revealed that three of the antibodies (1C6, P2-7E and P2-8G) recognized conserved antigen epitopes on capsid protein VP2, and exhibited broad reactivity against both FMDV serotypes A and O. An alanine-substitution scanning assay and sequence conservation analysis elucidated that these porcine mAbs recognized two conserved epitopes on VP2: a linear epitope (2KKTEETTLL10) in the N terminus and a conformational epitope involving residues K63, H65, L66, F67, D68 and L81 on two β-sheets (B-sheet and C-sheet) that depended on the integrity of VP2. Random parings of heavy and light chains of the IgGs confirmed that the heavy chain is predominantly involved in binding to antigen. The light chain of porcine IgG contributes to the binding affinity toward an antigen and may function as a support platform for antibody stability. In summary, this study is the first to reveal the conserved antigenic profile of FMDV recognized by porcine B cells and provides a novel method for analysing the antibody response against FMDV in its natural hosts (i.e. pigs) at the clonal level.

中文翻译：

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猪单克隆抗体的分离和表征揭示了口蹄疫病毒 VP2 上两个不同的血清型独立表位

猪易感染口蹄疫病毒（FMDV），体液免疫反应在预防口蹄疫病毒感染中起着至关重要的作用。然而，关于源自猪的单个 B 细胞的 FMDV 特异性 mAb 的信息很少。本研究旨在确定被猪抗体识别的 FMDV 的抗原特征。因此，使用基于荧光的单 B 细胞抗体技术开发了一组猪源性抗 FMDV 单克隆抗体。蛋白质印迹显示，三种抗体（1C6、P2-7E 和 P2-8G）识别衣壳蛋白 VP2 上的保守抗原表位，并对 FMDV 血清型 A 和 O 表现出广泛的反应性。丙氨酸替代扫描测定和序列保守性分析阐明这些猪单克隆抗体识别 VP2 上的两个保守表位：2 KKTEETTLL 10 ) 在 N 末端和一个构象表位，涉及两个 β 折叠（B 折叠和 C 折叠）上的残基 K63、H65、L66、F67、D68 和 L81，这取决于 VP2 的完整性。IgG 重链和轻链的随机配对证实重链主要参与与抗原的结合。猪 IgG 的轻链有助于对抗原的结合亲和力，并可作为抗体稳定性的支持平台。总之，这项研究首次揭示了猪 B 细胞识别的 FMDV 的保守抗原谱，并提供了一种在克隆水平上分析其天然宿主（即猪）中针对 FMDV 的抗体反应的新方法。