Key Points Four human Der p 2–specific IgE mAbs were identified by a new hybridoma technology. The human IgE mAbs recognize two nonoverlapping epitopes on Der p 2. NMR identified residues in the epitopes of human IgE and murine IgG mAbs. IgE Abs drive the symptoms of allergic disease upon cross-linking allergens on mast cells or basophils. If the IgE binding sites on the allergens could be identified, it may be useful for creating new forms of immunotherapy. However, direct knowledge of the human IgE (hIgE) epitopes is limited because of the very low frequency of IgE-producing B cells in blood. A new hybridoma technology using human B cells from house dust mite–allergic patients was used to identify four Der p 2–specific hIgE mAbs. Their relative binding sites were assessed and compared by immunoassays with three previously studied murine IgG mAbs. Immunoassays showed that the recognition of Der p 2 by the first three hIgE was inhibited by a single murine IgG, but the fourth hIgE recognized a different epitope from all the other mAbs. The functional ability of the hIgE that bind different epitopes to cross-link Der p 2 was demonstrated in a mouse model of passive systemic anaphylaxis. Nuclear magnetic resonance analyses of Der p 2 in complex with IgG and IgE Abs were used to identify specific residues in the epitopes. To our knowledge, the combination of immunoassays to distinguish overlapping epitopes and nuclear magnetic resonance analyses to identify specific residues involved in Ab binding provided the first epitope mapping of hIgE mAbs to an allergen. The technologies developed in this study will be useful in high-resolution mapping of human epitopes on other Ags and the design of improved therapeutics.

中文翻译：

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在主要尘螨过敏原 Der p 2 上绘制人类单克隆 IgE 表位

关键点 通过一种新的杂交瘤技术鉴定了四种人类 Der p 2 特异性 IgE mAb。人 IgE mAb 识别 Der p 2 上的两个非重叠表位。NMR 鉴定了人 IgE 和鼠 IgG mAb 表位中的残基。IgE Abs 在肥大细胞或嗜碱性粒细胞上交联过敏原后会引发过敏性疾病的症状。如果可以识别过敏原上的 IgE 结合位点，它可能有助于创造新的免疫疗法形式。然而，由于血液中产生 IgE 的 B 细胞的频率非常低，对人类 IgE (hIgE) 表位的直接了解是有限的。使用来自屋尘螨过敏患者的人类 B 细胞的新杂交瘤技术被用于鉴定四种 Der p 2 特异性 hIgE mAb。它们的相对结合位点通过免疫测定与三种先前研究的鼠 IgG mAb 进行评估和比较。免疫测定表明，前三个 hIgE 对 Der p 2 的识别被单个鼠 IgG 抑制，但第四个 hIgE 识别的表位与所有其他 mAb 不同。在被动全身性过敏反应的小鼠模型中证明了结合不同表位以交联 Der p 2 的 hIgE 的功能能力。与 IgG 和 IgE Abs 复合的 Der p 2 的核磁共振分析用于鉴定表位中的特定残基。据我们所知，区分重叠表位的免疫测定和鉴定与抗体结合所涉及的特定残基的核磁共振分析相结合，首次将 hIgE mAb 表位定位到过敏原。