Mechanistic understanding of fouling of protein A chromatography resin
Mechanistic understanding of fouling of protein A chromatography resin
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Protein A 层析树脂污染机理的理解
DOI：
10.1016/j.chroma.2016.06.084
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发表时间：
2016
影响因子：
4.1
通讯作者：
Pathak M
中科院分区：
文献类型：
--
作者：
Pathak M
本文旨在提供一个透彻的了解如何结垢的蛋白A树脂发生。已经检查了广泛使用的基于琼脂糖的蛋白A树脂MabSelect SuRe™的结合和质量传递特性，以了解树脂结垢的机制。可能存在影响树脂结垢的各种因素。这些包括由于进料中的组分导致的产物/杂质积聚和由于使用苛刻缓冲液导致的配体降解。为了阐明其贡献，在有和没有产品负载的情况下进行了循环研究。本文中给出的结果提供了对限制蛋白A色谱树脂寿命的原因的清晰理解。发现在第100次循环结束时，由于使用进料材料而导致的蛋白A树脂的容量下降是不使用进料材料的5倍。与新鲜树脂相比，循环的树脂样品显示颗粒孔隙率降低24%，孔传质系数降低51%。透射电子显微镜（TEM）用于定性监测污垢在循环树脂上的积累。结垢的树脂样品在树脂颗粒的内部和外部都含有致密的残留物，既作为珠表面处的膜又作为颗粒。表面活化能增加五倍的情况下，污垢的树脂样品。结垢的主要事件被确定为进料组分在树脂上的非特异性吸附，表明孔扩散是速率限制步骤。预计这些发现将有助于开发更稳健、更经济的单克隆抗体纯化下游生产工艺。
This paper aims to provide a thorough understanding of how fouling of Protein A resin takes place. Binding and mass transport properties of widely used agarose-based Protein A resin, MabSelect SuRe™, have been examined to understand the mechanism of resin fouling. There could be various factors that impact resin fouling. These include product/impurity build-up due to components in the feed material and ligand degradation due to the use of harsh buffers. To unravel their contributions, cycling studies were performed with and without product loading. The results presented in this paper provide a lucid understanding of the causative factors that limit Protein A chromatographic resin lifetime. The capacity fall for protein A resin at the end of 100th cycle due to use of feed material was found to be five times greater than that without using feed material. Compared to the fresh resin, the cycled resin samples shows 24% reduction in particle porosity and 51% reduction in pore mass transfer coefficient. Transmission electron microscopy (TEM) was used to qualitatively monitor accumulation of foulants on the cycled resin. Fouled resin sample contained a dense residue in the interior and exterior of resin particle both as a film at the bead surface and as granules. The surface activation energy increased five times in the case of fouled resin sample. The major event in fouling was identified as the non-specific adsorption of the feed material components on resin, signaling that pore diffusion is the rate limiting step. It is anticipated that these findings will assist in development of a more robust and economical downstream manufacturing process for monoclonal antibody purification.