Isolating recombinant antibodies against specific protein morphologies using atomic force microscopy and phage display technologies
Isolating recombinant antibodies against specific protein morphologies using atomic force microscopy and phage display technologies
复制标题
DOI：
10.1093/protein/gzl036
复制
发表时间：
2006-11-01
影响因子：
2.4
通讯作者：
Sierks, Michael R.
中科院分区：
文献类型：
--
作者：
Barkhordarian, Hedieh;Emadi, Sharareh;Sierks, Michael R.
分离不稳定、存在多种形态或可获得性非常有限的抗原的抗体可能非常困难。在这里，我们描述了一种新的技术，它结合了噬菌体展示抗体技术和原子力显微镜(AFM)的能力，用于分离与特定形态的靶抗原-α-突触核蛋白结合的抗体片段。原子力显微镜成像使我们既可以可视化目标抗原的存在和形态，也可以监测生物淘洗过程中每一步的效率。我们证明，仅经过两轮选择，就可以分离出针对靶抗原形态的噬菌体展示抗体。靶抗原α-突触核蛋白与帕金森氏病(PD)相关。α-突触核蛋白纤维聚集成路易体包涵体是帕金森病的一个显著特征。虽然α-突触核蛋白可以形成几种不同的聚集形态，包括低聚物、原纤维和纤维，但这些形态在帕金森病进展中的作用尚不清楚。重组抗体的成功选择具有潜在的治疗价值，因为单链片段变量(ScFv)可以在细胞内表达，以控制特定蛋白聚集体的折叠和毒性。
Isolation of antibodies to antigens that are either unstable, exist in multiple morphologies or have very limited availability can be prohibitively difficult. Here we describe a novel technique combining the capabilities of phage display antibody technology and atomic force microscopy (AFM) that is used to isolate antibody fragments that bind to a specific morphology of the target antigen, alpha-synuclein. AFM imaging allows us to both visualize the presence and morphology of the target antigen as well as to monitor the efficiency of each step in the bio-panning process. We demonstrate that phage displayed antibodies specific to the target antigen morphology can be isolated after only two rounds of selection. The target antigen, alpha-synuclein, has been correlated with the Parkinson's disease (PD). Accumulation of alpha-synuclein fibrillar aggregates into Lewy body inclusions is a hallmark feature of PD. While alpha-synuclein can form several different aggregate morphologies including oligomers, protofibrils and fibrils, the role of these morphologies in the progression of PD is not known. The successful selection of the recombinant antibody described here can have potential therapeutic value since the single-chain fragment variable (scFv) can be expressed intracellularly to control folding and toxicity of the specific protein aggregates.