Activation of Resin with Controllable Ligand Density via Catalytic Oxa-Michael Addition and Application in Antibody Purification
Activation of Resin with Controllable Ligand Density via Catalytic Oxa-Michael Addition and Application in Antibody Purification
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催化 Oxa-Michael 加成活化可控配体密度的树脂及其在抗体纯化中的应用
DOI：
10.1016/j.chroma.2018.07.032
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发表时间：
2018
影响因子：
4.1
通讯作者：
Qu Jingping
中科院分区：
文献类型：
--
作者：
Cheng Fang;Li Mingyang;He Wei;Sun Bingbing;Qin Jinyan;Qu Jingping
本文报道了一种以二乙烯基砜为原料，采用催化氧化-迈克尔加成反应可控活化树脂的新方法。通过对多种有机催化剂的筛选，发现PPh 3和DMAP在非质子溶液中具有较高的催化效率。X射线光电子能谱（XPS）分析表明，与传统的水相反应相比，反应效率高，副反应少，活化密度高。以PPh 3为催化剂，在12 h内，最大活化密度达到157.5 ± 1.2 μmol/g树脂，是传统水相反应的1.5倍。随后与色谱配体缀合，即，4-在使用巯基乙基吡啶（MEP）的情况下，树脂能够进行抗体纯化。以IgG和BSA为模型蛋白，研究了树脂的吸附等温线和动态结合行为。在具有较高配体密度的树脂上观察到IgG的较高亲和力和动态结合能力。最后，将树脂样品应用于从细胞培养上清液纯化治疗性单克隆抗体。具有高配体密度的树脂样品的回收率比商业树脂（MEP HyperCel）的回收率高70%。此外，我们的方法通过改变反应时间实现了可控的色谱配体密度，这有助于澄清密度-亲和力关系并改善工艺规模的抗体纯化。
This article reported a new strategy for resin activation with divinyl sulfone using catalytic oxa-Michael addition in a controllable manner. By screening a variety of organocatalysts, PPh3and DMAP stand out with high catalytic efficiency in aprotic solutions. X-ray photoelectron spectroscopy (XPS) analysis indicates high reaction efficiency and less side reactions than traditional aqueous reactions, resulting in high activation density. A maximum activation density of 157.5 ± 1.2 μmol/g resin was achieved in 12 h using PPh3as catalyst, which is 1.5 times higher than the traditional aqueous reactions. Followed by conjugation with a chromatographic ligand,i.e., 4-mercaptoethyl pyridine (MEP), the resin is capable of antibody purification. Using IgG and BSA as model proteins, adsorption isotherms and dynamic binding behavior of the resin samples were investigated. A higher affinity and dynamic binding capacity of IgG was observed on resins with higher ligand density. Finally, the resin samples were applied to the purification of a therapeutic monoclonal antibody from cell culture supernatant. The recovery of the resin samples with high ligand density are 70% higher than those of the commercial resin (MEP HyperCel). Moreover, our method achieves a controllable chromatographic ligand density by varying reaction times, which is useful to clarify the density-affinity relationship and improve process-scale antibody purification.