Purification of monoclonal antibodies from clarified cell culture fluid using Protein A capture continuous countercurrent tangential chromatography.
Purification of monoclonal antibodies from clarified cell culture fluid using Protein A capture continuous countercurrent tangential chromatography.
复制标题
使用蛋白质A捕获连续的逆时性切线色谱法从澄清的细胞培养液中纯化单克隆抗体。
DOI：
10.1016/j.jbiotec.2015.02.026
复制
发表时间：
2015-11-10
影响因子：
4.1
通讯作者：
Shinkazh O
中科院分区：
文献类型：
--
作者：
Dutta AK;Tran T;Napadensky B;Teella A;Brookhart G;Ropp PA;Zhang AW;Tustian AD;Zydney AL;Shinkazh O
最近使用简单模型系统的研究表明，连续逆流切向色谱（CCTC）有潜力克服使用填充柱的传统蛋白A色谱的许多局限性。本工作的目的是优化和实现CCTC系统纯化澄清中国仓鼠卵巢（CHO）细胞培养液的单克隆抗体使用商业蛋白a树脂。对之前的CCTC系统进行了一些改进，包括使用保留泵来保持流动浆体中的树脂浓度稳定，消除浆体储罐以提高生产率，并在结合步骤中引入“后粘合剂”以提高抗体回收率。建立了一种动力学结合模型，以估计多阶段结合步骤所需的停留时间，以优化收率和生产率。数据是通过纯化来自两个不同厂家的两种商业抗体获得的，一种是低滴度（~0.67 g/L），一种是高滴度（~6.9 g/L），证明了CCTC系统的通用性。宿主细胞蛋白去除率、抗体得率和纯度与常规柱层析法相似；然而，CCTC系统的生产率要高得多。这些结果清楚地证明了连续逆流切向色谱法用于单克隆抗体产品的商业纯化的能力。
Recent studies using simple model systems have demonstrated that Continuous Countercurrent Tangential Chromatography (CCTC) has the potential to overcome many of the limitations of conventional Protein A chromatography using packed columns. The objective of this work was to optimize and implement a CCTC system for monoclonal antibody purification from clarified Chinese Hamster Ovary (CHO) cell culture fluid using a commercial Protein A resin. Several improvements were introduced to the previous CCTC system including the use of retentate pumps to maintain stable resin concentrations in the flowing slurry, the elimination of a slurry holding tank to improve productivity, and the introduction of an “after binder” to the binding step to increase antibody recovery. A kinetic binding model was developed to estimate the required residence times in the multi-stage binding step to optimize yield and productivity. Data were obtained by purifying two commercial antibodies from two different manufactures, one with low titer (~0.67 g/L) and one with high titer (~6.9 g/L), demonstrating the versatility of the CCTC system. Host cell protein removal, antibody yields and purities were similar to that obtained with conventional column chromatography; however, the CCTC system showed much higher productivity. These results clearly demonstrate the capabilities of continuous countercurrent tangential chromatography for the commercial purification of monoclonal antibody products.