Novel strategy for selection of monoclonal antibodies against highly conserved antigens: phage library panning against ephrin-B2 displayed on yeast.
Novel strategy for selection of monoclonal antibodies against highly conserved antigens: phage library panning against ephrin-B2 displayed on yeast.
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针对高度保守的抗原选择单克隆抗体的新型策略：噬菌体库围绕酵母上显示的ephrin-b2。
DOI：
10.1371/journal.pone.0030680
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发表时间：
2012
期刊：
影响因子：
3.7
通讯作者：
Jin MM
中科院分区：
文献类型：
--
作者：
Gu X;Vedvyas Y;Chen X;Kaushik T;Hwang CI;Hu X;Nikitin AY;Jin MM
Ephrin-B2主要在动脉来源的内皮中表达，通过其与EphB 4的相互作用参与发育性血管生成和新血管形成。尽管肝配蛋白-B2在生理和病理条件中的重要性，但由于其在整个人类和啮齿动物中的序列高度保守性，因此产生针对肝配蛋白-B2的单克隆抗体一直具有挑战性。利用一种新的抗体筛选方法，通过淘选酵母中展示的抗原的人抗体噬菌体文库，我们已经分离出高亲和力的抗肝配蛋白-B2的抗体。一种高亲和力结合剂（命名为“EC 8”）的功能表现在其抑制肝配蛋白-B2与EphB 4相互作用、与鼠肝配蛋白-B2交叉反应以及诱导内化到表达肝配蛋白-B2的细胞中的能力。EC 8也与免疫沉淀和标准化学固定程序后检测组织中ephrin-B2表达相容。与先前关于在一些上皮肿瘤和肿瘤相关血管中肝配蛋白-B2诱导的报道一致，EC 8特异性地检测肿瘤以及肿瘤内外的血管中的肝配蛋白-B2。我们设想，在这项研究中开发的单克隆抗体可用作一种试剂，以探测ephrin-B2在正常以及病理条件下的分布，并拮抗ephrin-B2与EphB 4的相互作用的基础科学和治疗应用。
Ephrin-B2 is predominately expressed in endothelium of arterial origin, involved in developmental angiogenesis and neovasculature formation through its interaction with EphB4. Despite its importance in physiology and pathological conditions, it has been challenging to produce monoclonal antibodies against ephrin-B2 due to its high conservation in sequence throughout human and rodents. Using a novel approach for antibody selection by panning a phage library of human antibody against antigens displayed in yeast, we have isolated high affinity antibodies against ephrin-B2. The function of one high affinity binder (named as ‘EC8’) was manifested in its ability to inhibit ephrin-B2 interaction with EphB4, to cross-react with murine ephrin-B2, and to induce internalization into ephrin-B2 expressing cells. EC8 was also compatible with immunoprecipitation and detection of ephrin-B2 expression in the tissue after standard chemical fixation procedure. Consistent with previous reports on ephrin-B2 induction in some epithelial tumors and tumor-associated vasculatures, EC8 specifically detected ephrin-B2 in tumors as well as the vasculature within and outside of the tumors. We envision that monoclonal antibody developed in this study may be used as a reagent to probe ephrin-B2 distribution in normal as well as in pathological conditions and to antagonize ephrin-B2 interaction with EphB4 for basic science and therapeutic applications.
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影响因子：
11.2
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