Porcine epidemic diarrhea virus (PEDV) infection causes an acute enteric tract infectious disease characterized by vomiting, anorexia, dehydration, weight loss and high mortality in neonatal piglets. During PEDV infection, the spike protein (S) is a major virion structural protein interacting with receptors and inducing neutralizing antibodies. However, the neutralizing B-cell epitopes within PEDV S protein have not been well studied. To accurately identify the important immunodominant region of S1, the purified truncated S1 proteins (SA, SB, SC, SD and SE) were used to immunize BALB/c mice to prepare polyclonal antibodies. The antisera titers were determined by indirect ELISA, western blot and IFA after four immunizations to find the important immunodominant region of S1, and then purified the immunodominant region of S1 protein and immunized mice to generate the special antibodies, and then used recombinant peptides to determine the B-cell epitopes of monoclonal antibodies. Five antisera of recombinant proteins of the spike protein region of PEDV were generated and we found that only the polyclonal antibody against part of the S1 region (signed as SE protein, residues 666–789) could recognize the native PEDV. Purified SE protein was used to immunize BALB/c mice and generate mAb 2E10. Pepscan of the SE protein demonstrated that SE16 (722SSTFNSTREL731) is the minimal linear epitope required for reactivity with the mAb 2E10. Further investigation indicated that the epitope SE16 was localized on the surface of PEDV S protein in the 3D structure. A mAb 2E10 that is specifically bound to PEDV was generated and identified a specific linear B-cell epitope (SE16, 722SSTFNSTREL731) of the mAb. The epitope region of PEDV S1 localized in the different regions in comparison with the earlier identified epitopes. These findings enhance the understanding of the PEDV spike protein structure for vaccine design and provide a potential use for developing diagnostic methods to detect PEDV.

中文翻译：

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在猪流行性腹泻病毒的刺突蛋白中鉴定出新的B细胞表位。

猪流行性腹泻病毒（PEDV）感染会导致急性肠道传染病，其特征是新生仔猪呕吐，厌食，脱水，体重减轻和高死亡率。在PEDV感染期间，刺突蛋白（S）是一种主要的病毒体结构蛋白，可与受体相互作用并诱导中和抗体。然而，PEDV S蛋白中的中和B细胞表位尚未得到很好的研究。为了准确识别S1的重要免疫优势区域，使用纯化的截短S1蛋白（SA，SB，SC，SD和SE）免疫BALB / c小鼠以制备多克隆抗体。四次免疫后，通过间接ELISA，western blot和IFA测定抗血清滴度，以发现S1的重要免疫优势区域，然后纯化S1蛋白的免疫优势区域并免疫小鼠以产生特殊抗体，然后使用重组肽确定单克隆抗体的B细胞表位。产生了五种PEDV刺突蛋白区域重组蛋白的抗血清，我们发现只有针对S1部分区域的多克隆抗体（标记为SE蛋白，残基666-789）可以识别天然PEDV。纯化的SE蛋白用于免疫BALB / c小鼠并产生mAb 2E10。SE蛋白的Pepscan证明SE16（722SSTFNSTREL731）是与mAb 2E10反应所需的最小线性表位。进一步的研究表明，表位SE16位于3D结构中的PEDV S蛋白的表面上。产生了与PEDV特异性结合的mAb 2E10，并鉴定了该mAb的特定线性B细胞表位（SE16、722SSTFNSTREL731）。与早期鉴定的表位相比，PEDV S1的表位区域位于不同的区域。这些发现增强了对用于疫苗设计的PEDV刺突蛋白结构的了解，并为开发检测PEDV的诊断方法提供了潜在用途。