PDF(Pubmed)
Abstract：
Antibodies (mAbs) and antibody fragments (Fabs) constitute one of the largest and most rapidly expanding groups of protein pharmaceuticals. In particular, antibody fragments have certain advantages over mAbs in some therapeutic settings. However, due to their greater chemical diversity, they are more challenging to purify for large-scale production using a standard purification platform. Besides, the removal of Fab-related byproducts poses a difficult purification challenge. Alternative Fab purification platforms could expedite their commercialization and reduce the cost and time invested. Accordingly, we employed a strong cation exchanger using a pH-based, highly linear gradient elution mode following Protein L affinity purification and developed a robust two-step purification platform for an antibody fragment. The optimized pH gradient elution conditions were determined on the basis of purity level, yield, and the abundance of Fab-related impurities, particularly free light chain. The purified Fab molecule Ranibizumab possessed a high degree of similarity to its originator Lucentis. The developed purification platform highly intensified the process and provided successful clearance of formulated Fab- and process-related impurities (∼98 %) with an overall process recovery of 50 % and, thus, might be a new option for Fab purification for both academic and industrial purposes.
摘要：
抗体(mAb)和抗体片段(Fab)构成最大和最快速扩展的蛋白质药物组之一。特别是,在一些治疗环境中,抗体片段比单克隆抗体具有某些优势.然而,由于它们更大的化学多样性，它们对于使用标准纯化平台进行大规模生产的纯化更具挑战性。此外,Fab相关副产物的去除带来了困难的纯化挑战。替代Fab纯化平台可以加快其商业化并降低成本和投入的时间。因此，我们使用了一种基于pH的强阳离子交换剂，蛋白L亲和纯化后的高度线性梯度洗脱模式，并为抗体片段开发了强大的两步纯化平台。基于纯度水平确定优化的pH梯度洗脱条件,产量，以及大量的Fab相关杂质，特别是自由轻链。纯化的Fab分子Ranibizumab与其鼻祖Lucentis具有高度的相似性。开发的纯化平台高度强化了该过程，并成功清除了配制的Fab和与过程相关的杂质（〜98％），总体过程回收率为50％，并且因此，可能是用于学术和工业目的的Fab纯化的新选择。
