应用于抗体药物捕获的耐碱亲和层析介质性能的比较

Comparison of performances of alkaline-resistant affinity chromatographic media for capture of therapeutic antibodies

目的比较应用于抗体药物捕获的耐碱的4种蛋白A亲和层析介质和2种小分子亲和层析介质的性能,为单抗纯化工艺的选择提供参考。方法利用两种单抗纯品（mAb1和mAb2）测定4种蛋白A亲和层析介质MabSelectSuRe、POROS MabCaptureA、Absolute High Cap、TOYOPEARL AF-rProtein A-650F和2种小分子亲和层析介质Mabsorbent A2P HF、Fabsorbent F1P HF在停留时间分别为4、6、8 min时的动态载量;利用含mAb2的发酵液,从回收率和杂质去除方面比较6种亲和层析介质的纯化效果。结果在5%穿透点,各介质对mAb1和mAb2的动态载量在35～73 g/L之间。小分子亲和层析介质Mabsorbent A2P HF纯化mAb2的回收率为89%,其他5种介质纯化mAb2的回收率均≥96%;6种介质纯化mAb2的宿主细胞蛋白（host cell protein,HCP）在2 000 ppm之内,蛋白A残留量在20 ppm之内。结论结合流速、动态载量、回收率和杂质去除效果等指标,可从6种亲和层析介质中挑选适用于抗体类药物下游纯化工艺的耐碱型蛋白A或小分子亲和介质。

Objective To compare the performances of four alkaline-resistant protein A affinity chromatographic media and two small molecular affinity chromatographic media,and provide a reference for selection of purification procedure for monoclonal antibodies（mAbs）. Methods The dynamic binding capacities（DBCs）of four protein A affinity chromatographic media MabSelectSuRe,POROS MabCaptureA,Absolute High Cap and TOYOPEARL AF-rProtein A-650F as well as two small molecular affinity chromatographic media Mabsorbent A2P HF and Fabsorbent F1P HF were determined by plotting breakthrough curves of two purified monoclonal antibodies（mAb1 and mAb2）with retention times of 4,6 and 8 min,respectively. The performance of these six affinity media,including recovery rate and removal of foreign matters,were further investigated with clarified cell culture supernatant containing mAb2. Results At 5% breakthrough point,the DBCs of these media were 35 ～ 73 g / L. The recovery rate of mAb2 by Mabsorbent A2P HF was 89%,while those by other five media were not less than 96%. The residual host cell protein（HCP）contents in mAb2 purified by the six media were less than 2 000 ppm,while the residual protein A contents were less than 20 ppm. Conclusion Taking into consideration of flow rate,DBC,recovery rate and removal of foreign matters,alkaline-resistant protein A or small molecular affinity chromatographic media suitable for downstream purification process of therapeutic antibodies may be selected from the six affinity chromatographic media.