Method development for the separation of monoclonal antibody charge variants in cation exchange chromatography, Part II: pH gradient approach
Method development for the separation of monoclonal antibody charge variants in cation exchange chromatography, Part II: pH gradient approach
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DOI：
10.1016/j.jpba.2014.09.032
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发表时间：
2015-01-05
影响因子：
3.4
通讯作者：
Guillarme, Davy
中科院分区：
文献类型：
--
作者：
Fekete, Szabolcs;Beck, Alain;Guillarme, Davy
对阳离子交换 pH 梯度方法进行了评估，以表征 10 种模型单克隆抗体，包括帕尼单抗、那他珠单抗、西妥昔单抗、贝伐单抗、曲妥珠单抗、利妥昔单抗、帕利珠单抗、阿达木单抗、狄诺塞麦和奥法木单抗。这项工作表明，仅基于四次初始运行（即两次梯度时间和两次流动相），可以在阳离子交换 pH 梯度模式下对保留和分离进行建模温度）。使用 100 mm x 4.6 mm 强阳离子交换柱时，仅需 6 小时即可完成完整的方法优化。预测的准确性非常好，预测保留时间与实验保留时间之间的平均差异约为1%。10种模型抗体在pH和盐梯度模式下均成功洗脱，证明两种洗脱模式都可以被认为是多产物电荷敏感分离方法。对于大多数化合物，在盐梯度模式下可以更好地解析变体，并且盐梯度方法中的峰容量也更高。这些观察结果证实，在抗体表征方面，pH 梯度法可能不如盐梯度阳离子交换色谱法更受关注。 (C) 2014 Elsevier B.V. 保留所有权利。
The cation exhange pH gradient approach was evaluated for the characterization of 10 model monoclonal antibodies including panitumumab, natalizumab, cetuximab, bevacizumab, trastuzumab, rituximab, palivizumab, adalimumab, denosumab and ofatumumab.This work shows that retention and resolution can be modelled in cation exchange pH gradient mode, based on only four initial runs (i.e. two gradient times and two mobile phase temperature). Only 6 h were required for a complete method optimization when using a 100 mm x 4.6 mm strong cation exchange column. The accuracy of the predictions was excellent, with an average difference between predicted and experimental retention times of about 1%.The 10 model antibodies were successfully eluted in both pH and salt gradient modes, proving that both modes of elution can be considered as multi-product charge sensitive separation methods. For most of the compounds, the variants were better resolved in the salt gradient mode and the peak capacities were also higher in the salt gradient approach. These observations confirm that pH gradient approach may be of lower interest than salt gradient cation exchange chromatography for antibody characterization. (C) 2014 Elsevier B.V. All rights reserved.