Determination of Protein Aggregation With Differential Mobility Analysis: Application to IgG Antibody
Determination of Protein Aggregation With Differential Mobility Analysis: Application to IgG Antibody
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DOI：
10.1002/bit.22017
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发表时间：
2008-12-15
影响因子：
3.8
通讯作者：
Tarlov, Michael J.
中科院分区：
文献类型：
--
作者：
Pease, Leonard F., III;Elliott, John T.;Tarlov, Michael J.
在这里，我们描述了使用电喷雾微分迁移率分析 (ES-DMA)（也称为 Os 相电泳迁移率分子分析）作为测量溶液中蛋白质低阶可溶性聚集体的方法。我们用 IgG 抗体证明了概念。在 ES-DMA 中，抗体蛋白的水溶液被电喷雾，并使用微分迁移率分析仪根据其电泳迁移率来分离各种雾化物质。通过这种方式，可以使用当前设置获得 3 至 250 nm 范围内蛋白质种类的完整尺寸分布，包括 IgG 单体到五聚体的不同峰。通过 DMA 测量的 IgG 和 IgG 聚集体的大小与通过简单模型计算得出的大小非常一致，简单模型从蛋白质晶体学数据中获取 IgG 的结构尺寸。还检查了 IgG 聚集对溶液浓度和离子强度的依赖性，并对含有化学交联抗体的聚集体部分进行了定量。这些结果表明，ES-DMA 具有作为测量工具的潜力，可用于研究蛋白质聚集现象，例如与抗体试剂制造和蛋白质治疗相关的现象。
Here we describe the use of electrospray differential mobility analysis (ES-DMA), also known a Os-phase electrophoretic mobility molecular analysis as a method for measuring low-order soluble aggregates of proteins in solution. We demonstrate proof of concept with IgG antibodies. In ES-DMA, aqueous Solutions of the antibody protein are electrosprayed and the various aerosolized species are separated according to their electrophoretic mobility using a differential mobility analyzer. In this way, complete size distributions of protein species present from 3 to 250 nm can be obtained with the current set up, including distinct peaks for IgG monomers to pentamers. The sizes of the IgG and IgG aggregates measured by DMA were found to be in good agreement with those calculated from simple models, which take the structural dimensions of IgG from protein crystallographic data. The dependence of IgG aggregation on the solution concentration and ionic strength was also examined, and the portion of aggregates containing chemically crosslinked antibodies was quantified. These results indicate that ES-DMA holds potential as as a measurement tool to study protein aggregation phenomena such as those associated with antibody reagent manufacturing and protein therapeutics.