Qu Ju, Wan Yan, Li Yifeng
Downstream Process Development, WuXi Biologics, 288 Fute Zhong Road, Waigaoqiao Free Trade Zone, Shanghai, 200131, China.
J Biol Methods. 2024 Jul 10;11(2):e99010016. doi: 10.14440/jbm.2024.0006. eCollection 2024.
Protein A and Protein L affinity chromatographies are extensively used in mAb and bispecific antibody (bsAb) purification. In addition to product capture, they are both capable of separating certain product-related by-products and aggregates under appropriate conditions. For both types of chromatography, previous studies suggested that adding a salt additive to the mobile phase can significantly improve the resolution between product and by-products/aggregates. Nevertheless, the effects of different salt additives on antibody elution in Protein A and Protein L chromatography have not been compared. In the current study, we compared the effects of three salt additives, sodium chloride (NaCl), calcium chloride (CaCl), and arginine hydrochloride (Arg·HCl), on antibody elution in Protein A and Protein L chromatography. Interestingly, while NaCl suppressed antibody elution in both types of chromatography, CaCl, and Arg·HCl promoted antibody elution in Protein A chromatography but suppressed antibody elution in Protein L chromatography. In addition, we evaluated the effect of each salt gradient on aggregate removal by Protein L chromatography. The information provided by the current study should be useful to the selection of conditions/additives for improving by-product removal by Protein A and Protein L chromatography.
蛋白A亲和层析和蛋白L亲和层析被广泛应用于单克隆抗体（mAb）和双特异性抗体（bsAb）的纯化。除了捕获产物外，它们在适当条件下都能够分离某些与产物相关的副产物和聚集体。对于这两种层析类型，先前的研究表明，在流动相中添加盐添加剂可以显著提高产物与副产物/聚集体之间的分离度。然而，不同盐添加剂对蛋白A和蛋白L层析中抗体洗脱的影响尚未进行比较。在本研究中，我们比较了三种盐添加剂，即氯化钠（NaCl）、氯化钙（CaCl）和盐酸精氨酸（Arg·HCl），对蛋白A和蛋白L层析中抗体洗脱的影响。有趣的是，虽然NaCl在两种层析类型中均抑制抗体洗脱，但CaCl和Arg·HCl在蛋白A层析中促进抗体洗脱，而在蛋白L层析中抑制抗体洗脱。此外，我们评估了每种盐梯度对蛋白L层析去除聚集体的效果。本研究提供的信息对于选择改善蛋白A和蛋白L层析去除副产物的条件/添加剂应是有用的。
