Combinatory antibody library display technologies have been invented and successfully implemented for the selection and engineering of therapeutic antibodies. Precise targeting of important epitopes on the protein of interest is essential for such isolated antibodies to serve as effective modulators of molecular interactions. We developed a strategy to efficiently isolate antibodies against a specific epitope on a target protein from a yeast display antibody library using dengue virus envelope protein domain III as a model target. A domain III mutant protein with a key mutation inside a cross-reactive neutralizing epitope was designed, expressed, and used in the competitive panning of a yeast display naïve antibody library. All the yeast display antibodies that bound to the wild type domain III but not to the mutant were selectively sorted and characterized. Two unique clones were identified and showed cross-reactive binding to envelope protein domain IIIs from different serotypes. Epitope mapping of one of the antibodies confirmed that its epitope overlapped with the intended neutralizing epitope. This novel approach has implications for many areas of research where the isolation of epitope-specific antibodies is desired, such as selecting antibodies against conserved epitope(s) of viral envelope proteins from a library containing high titer, high affinity non-neutralizing antibodies, and targeting unique epitopes on cancer-related proteins.

已经发明并成功实施了组合抗体库展示技术，用于治疗性抗体的选择和工程化。精确靶向目标蛋白质上的重要表位对于此类分离的抗体作为分子相互作用的有效调节剂至关重要。我们开发了一种策略，以使用登革病毒包膜蛋白结构域 III 作为模型靶标，从酵母展示抗体库中有效分离针对靶蛋白上特定表位的抗体。在交叉反应中和表位内具有关键突变的域 III 突变蛋白被设计、表达并用于酵母展示幼稚抗体库的竞争性淘选。所有与野生型结构域 III 结合但不与突变体结合的酵母展示抗体都被选择性地分类和表征。鉴定了两个独特的克隆，并显示出与来自不同血清型的包膜蛋白结构域 III 的交叉反应性结合。抗体之一的表位作图证实其表位与预期的中和表位重叠。这种新方法对需要分离表位特异性抗体的许多研究领域都有影响，例如从包含高滴度、高亲和力非中和抗体的文库中选择针对病毒包膜蛋白保守表位的抗体，以及靶向癌症相关蛋白上的独特表位。鉴定了两个独特的克隆，并显示出与来自不同血清型的包膜蛋白结构域 III 的交叉反应性结合。抗体之一的表位作图证实其表位与预期的中和表位重叠。这种新方法对需要分离表位特异性抗体的许多研究领域都有影响，例如从包含高滴度、高亲和力非中和抗体的文库中选择针对病毒包膜蛋白保守表位的抗体，以及靶向癌症相关蛋白上的独特表位。鉴定了两个独特的克隆，并显示出与来自不同血清型的包膜蛋白结构域 III 的交叉反应性结合。抗体之一的表位作图证实其表位与预期的中和表位重叠。这种新方法对需要分离表位特异性抗体的许多研究领域都有影响，例如从包含高滴度、高亲和力非中和抗体的文库中选择针对病毒包膜蛋白保守表位的抗体，以及靶向癌症相关蛋白上的独特表位。