Abstract：
A design procedure for the separation of charge variants of a monoclonal antibody (mAb) was developed, which was based on the coupling of cation-exchange chromatography (CEX) and anion-exchange chromatography (AEX) under high loading conditions. The design of the coupled process was supported by a dynamic model. The model was calibrated on the basis of band profiles of variants determined experimentally for the mAb materials of different variant compositions. The numerical simulations were used to select the coupling configuration and the loading conditions that allowed for efficient separation of the mAb materials into three products enriched with each individual variant: the acidic (av), main (mv) and basic (bv) one. In the CEX section, a two-step pH gradient was used to split the loaded mass of mAb into a weakly bound fraction enriched with av and mv, and a strongly bound fraction containing the bv-rich product. The weakly bound fraction was further processed in the AEX section, where the mv-rich product was eluted in flowthrough, while the av-rich product was collected by a step change in pH. The choice of flow distribution and the number of columns in the CEX and AEX sections depended on the variant composition of the mAb material. For the selected configurations, the optimized mAb loading density in the CEX columns ranged from 10 to 26 mg mL-1, while in the AEX columns it was as high as 300 or 600 mg mL-1, depending on the variant composition of the mAb material. By proper selection of the loading condition, a trade-off between yield and purity of the products could be reached.
摘要：
开发了用于分离单克隆抗体（mAb）的电荷变体的设计程序，其基于在高负载条件下的阳离子交换色谱(CEX)和阴离子交换色谱(AEX)的偶联。耦合过程的设计由动态模型支持。基于针对不同变体组成的mAb材料实验确定的变体的条带谱来校准模型。使用数值模拟来选择偶联构型和负载条件，这些条件允许将mAb材料有效分离成富含每种单独变体的三种产物：酸性（av），主要（MV）和基本（BV）之一。在CEX部分，使用两步pH梯度将mAb的负载质量分成富含av和mv的弱结合级分，和含有富含bv的产品的强结合级分。弱结合部分在AEX部分进一步处理，富含mv的产物在流过液中洗脱，同时通过pH的阶跃变化收集富含av的产品。在CEX和AEX部分中的流动分布和柱数的选择取决于mAb材料的变体组成。对于选定的配置，CEX柱中优化的mAb装载密度范围为10至26mgmL-1,而在AEX柱中,它高达300或600mgmL-1,这取决于mAb材料的变体组成。通过适当选择加载条件，可以达到产品的产率和纯度之间的权衡。
