对柱子负载和通流的SEC-HPLC分析提供了对低蛋白质A阶段收益率的批判性理解
Chen Liu1, Mengying Tian1, Wanyuan Dong1
1Downstream Process Development (DSPD), WuXi Biologics, 288 Fute Zhong Road, Waigaoqiao Free Trade Zone, Shanghai, 200131, China.
Protein expression and purification
|December 23, 2023
概括
细胞培养中的非结合聚合物导致抗体和Fc融合蛋白质的蛋白质A染色学产量较低. 建议对蛋白A负载和流通分析进行大小排除色谱-高性能液态色谱 (SEC-HPLC) 分析,以识别这些聚合物.
科学领域:
- 生物制药制造业 生物制药制造业
- 蛋白质染色学 蛋白质染色学
- 过程开发 过程开发
背景情况:
- 低蛋白A捕获产量 (约80%) 经常观察到单克隆抗体 (mAbs),双特异性抗体 (bsAbs) 和Fc融合蛋白.
- 之前的研究表明,细胞培养过程中形成的非结合聚合物是减少蛋白A阶段产量的潜在根本原因.
研究的目的:
- 在特定的生物制药制造项目中调查和确认非结合性聚合物的存在是低蛋白A产量的原因.
- 阐明这些聚合物导致蛋白A捕获效率降低的机制.
主要方法:
- 选择了5个生物制药项目,这些项目具有较低的A蛋白产量.
- 使用大小排除染色法-高性能液态染色法 (SEC-HPLC) 分析蛋白质A染色学负荷和流量样本.
主要成果:
- 通过SEC-HPLC,在所有选定的项目中,在蛋白A负载和流量样本中始终检测到非约束性聚合物.
- 这些聚合物的大尺寸被确定为它们无法与蛋白质A树脂结合的主要原因,因为它们无法穿透树脂珠.
结论:
- 蛋白A负载和流通的SEC-HPLC分析,虽然不是标准,但提供了对意想不到的蛋白A染色学性能的关键见解.
- 这种分析方法强烈推用于故障排除和优化抗体和Fc融合蛋白质的蛋白质A捕获过程,由于聚合物形成,其产量较低.
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