METHODS: Epitope mapping of RmS-17 was achieved using in silico approach combining linear B-cell epitope and antigenicity predictor. In addition, epitope mapping using overlapping peptides in an ELISA screening was used. The serpin tridimensional structure and the epitopes spatial location within the molecule were determined. Peptides were synthetized based on the predictions and used for the production of rabbit anti-sera. Purified IgG\'s were used to assess the antibodies capacity to neutralize RmS-17.
RESULTS: Through in silico mapping, nine potential B cell epitope regions were screened, with p1RmS-17 and p2RmS-17 selected for the experiment based on antigen prediction. In the ELISA screening using overlapping peptides, eight antibody-binding regions were identified, and p3RmS-17 and p4RmS-17 were chosen. Antibodies raised against p3RmS-17 and p4RmS-17 partially neutralized RmS-17 activity.
CONCLUSIONS: It was found that antibodies against a single epitope are sufficient to partially neutralize RmS-17 activity. These findings support the possibility of using an epitope-based vaccine for immunization against R. microplus.
方法:使用结合线性B细胞表位和抗原性预测因子的计算机模拟方法实现RmS-17的表位定位。此外,使用在ELISA筛选中使用重叠肽的表位作图。确定了serpin三维结构和分子内的表位空间位置。基于预测合成肽并用于生产兔抗血清。纯化的IgG用于评估抗体中和RmS-17的能力。
结果:通过计算机绘图,筛选了9个潜在的B细胞表位区域,根据抗原预测选择p1RmS-17和p2RmS-17进行实验。在使用重叠肽的ELISA筛选中,鉴定了8个抗体结合区,选择p3RmS-17和p4RmS-17。针对p3RmS-17和p4RmS-17产生的抗体部分中和RmS-17活性。
结论:发现针对单个表位的抗体足以部分中和RmS-17活性。这些发现支持使用基于表位的疫苗免疫针对R.microplus的可能性。