Conformation-specific antibodies against multiple amyloid protofibril species from a single amyloid immunogen
Conformation-specific antibodies against multiple amyloid protofibril species from a single amyloid immunogen
复制标题
DOI：
10.1111/jcmm.14119
复制
发表时间：
2019-03-01
影响因子：
5.3
通讯作者：
Graham, W. Vallen
中科院分区：
文献类型：
--
作者：
Bonito-Oliva, Alessandra;Schedin-Weiss, Sophia;Graham, W. Vallen
我们设计并使用了一种伴侣样淀粉样蛋白结合蛋白核结合蛋白1 （NUCB1）来稳定人胰岛淀粉样多肽（hIAPP）原原纤维，用于小鼠的免疫原。我们获得了多个对hIAPP原纤维有反应的单克隆抗体（mAb）克隆。进行了二次筛选，以确定与淀粉样蛋白β肽（A β 42）原原纤维交叉反应的克隆，但不与A β 40单体交叉反应。这些单抗在几项体外实验、阿尔茨海默病（AD）小鼠模型的免疫组织学研究和AD患者脑组织中得到了进一步的表征。我们发现用NUCB1-hIAPP复合物免疫小鼠获得的单克隆抗体与A β 42交叉反应，特异性靶向原原纤维并抑制其进一步聚集。与构象特异性结合一致，单抗似乎与病变组织中的细胞内抗原反应，但不与淀粉样斑块反应。我们假设我们在这里描述的单克隆抗体识别多个淀粉样原纤维共同的二级或四级结构表位。总之，我们报告了一种方法来创建单克隆抗体是构象敏感和序列无关的，可以针对多种类型的原纤维物种。
We engineered and employed a chaperone-like amyloid-binding protein Nucleobindin 1 (NUCB1) to stabilize human islet amyloid polypeptide (hIAPP) protofibrils for use as immunogen in mice. We obtained multiple monoclonal antibody (mAb) clones that were reactive against hIAPP protofibrils. A secondary screen was carried out to identify clones that cross-reacted with amyloid beta-peptide (A beta 42) protofibrils, but not with A beta 40 monomers. These mAbs were further characterized in several in vitro assays, in immunohistological studies of a mouse model of Alzheimer's disease (AD) and in AD patient brain tissue. We show that mAbs obtained by immunizing mice with the NUCB1-hIAPP complex cross-react with A beta 42, specifically targeting protofibrils and inhibiting their further aggregation. In line with conformation-specific binding, the mAbs appear to react with an intracellular antigen in diseased tissue, but not with amyloid plaques. We hypothesize that the mAbs we describe here recognize a secondary or quaternary structural epitope that is common to multiple amyloid protofibrils. In summary, we report a method to create mAbs that are conformation-sensitive and sequence-independent and can target more than one type of protofibril species.