人免疫球蛋白G酶解制备Fab和Fc片段及其分离纯化

Preparation and separation of Fab and Fc fragments from human immunoglobulin G with papain digestion

为了制备抗体Fab和Fc片段,采用木瓜蛋白酶酶解人免疫球蛋白G(IgG),通过优化酶解条件和色谱法分离过程,得到了纯度较高的Fab片段和Fc片段。考察了酶解pH、酶加入量、添加半胱氨酸和酶解时间等对IgG酶解过程的影响,优化了酶解条件,提高酶解效率,IgG转化率大于98%。酶解产物通过Protein A亲和色谱法和DEAE阴离子交换色谱法进行了纯化,分离得到Fc片段和Fab片段,收率分别为72.6%和40.1%。经SEC-HPLC分析,Fc片段纯度达95.7%,Fab片段纯度达96.6%。

For the preparation of Fab and Fc fragments with high purity,human immunoglobulin G（hIgG） was treated with papain and its purification with chromatographic techniques was explored.The concentration of cystein,papain addition,pH and incubation time were optimized to improve digestion efficiency with papain,and conversion of IgG could reach more than 98%.Then Protein A affinity chromatography and DEAE anion-exchange chromatography were used to separate the Fab and Fc fragments from the digestion mixture of hIgG.Based on the SEC-HPLC analysis,the purity of Fab fragment was 96.6% with the yield of 40.1%,while the purity of Fc fragment was 95.7% with the yield of 72.6%.