Efficient generation of human IgA monoclonal antibodies
Efficient generation of human IgA monoclonal antibodies
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DOI：
10.1016/j.jim.2015.04.010
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发表时间：
2015-07-01
影响因子：
2.2
通讯作者：
Mouquet, Hugo
中科院分区：
文献类型：
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作者：
Lorin, Valerie;Mouquet, Hugo
免疫球蛋白A（伊加）是人类产生的最丰富的抗体同种型。伊加抗体主要确保粘膜表面对入侵病原体的免疫保护，但也循环并大量存在于血液中。IgA在分子水平上是异质的，具有两种伊加亚型和通过与连接的α）链相互作用形成多聚体的能力。在这里，我们已经开发了一种有效的策略，以快速生成人的单克隆抗体的单体和二聚体形式的IgA 1和IgA 2。在表达克隆和瞬时转染293-F细胞后，大量产生了靶向HIV-1包膜蛋白的原型IgG 1单克隆抗体10-1074的重组单体和二聚体IgA 1/IgA 2对应物。使用接枝有抗IgA嵌合Fab的新型基于亲和力的树脂对10-1074 IgA进行HPLC纯化，然后使用基于尺寸排阻的FPLC ELISA结合实验进行单体/多聚体分离，证实10-1074的人工伊加类别转换不改变其抗原识别。总之，我们的技术方法允许非常有效地生产各种形式的纯化的重组人伊加分子，其是解剖生理和病理生理条件下的伊加B细胞应答以及研究IgA的生物学、功能和治疗潜力的宝贵工具。(C)2015爱思唯尔B. V.保留所有权利。
Immunoglobulin A (IgA) is the most abundant antibody isotype produced in humans. IgA antibodies primarily ensure immune protection of mucosal surfaces against invading pathogens, but also circulate and are present in large quantities in blood. IgAs are heterogeneous at a molecular level, with two IgA subtypes and the capacity to form multimers by interacting with the joining a) chain. Here, we have developed an efficient strategy to rapidly generate human IgA1 and IgA2 monoclonal antibodies in their monomeric and dimeric forms. Recombinant monomeric and dimeric IgA1/IgA2 counterparts of a prototypical IgG1 monoclonal antibody, 10-1074, targeting the HIV-1 envelope protein, were produced in large amounts after expression cloning and transient transfection of 293-F cells. 10-1074 IgAs were FPLC-purified using a novel affinity-based resin engrafted with anti-IgA chimeric Fabs, followed by a monomers/multimers separation using size exclusion-based FPLC ELISA binding experiments confirmed that the artificial IgA class switching of 10-1074 did not alter its antigen recognition. In summary, our technical approach allows the very efficient production of various forms of purified recombinant human IgA molecules, which are precious tools in dissecting IgA B-cell responses in physiological and pathophysiological conditions, and studying the biology, function and therapeutic potential of IgAs. (C) 2015 Elsevier B.V. All rights reserved.