间臂和丁基密度对疏水层析分离纯化重组乙肝病毒表面抗原的影响

Comparison of Adsorbent with Varying Arm Length and Ligand Density for the Purification of Recombinant Hepatitis B Virus Surface Antigen

摘要:

以国产高交联度的快流速琼脂糖为基质，合成了针对纯化中国仓鼠卵巢细胞表达乙肝病毒表面抗原（CHO-HBsAg）的不同间臂（3C、8C和10C）和不同配基密度的丁基疏水介质。配基密度的增加和间臂C链的延长都会增加介质的疏水性，从而影响其对CHO-HBsAg的分离效果。采用正交实验考察了配基密度、盐浓度、pH值三个影响因素对不同间臂疏水介质性能的影响。以分离CHO表达的乙肝病毒表面抗原（CHO-HBsAg）的收率和纯化倍数为主要指标。根据正交试验结果，分离效果最佳的介质间臂为8C、配基密度为22μmol/mL。该介质在硫酸铵浓度9%、pH7.0条件下，CHO-HBsAg的收率可以接近100%，纯化倍数达到60。

Abstract:

Novel hydrophobic absorbents were synthesized by immobilizing butyl derivative onto the highly cross-linked agarose beads manufatured in China, which are used as matrix. The effect of the spacer arm length (3C, 8C and 10C) and ligand density (from 13 to 45μmol/mL) on the hydrophobicity were investigated using purified Hepatitis B surface antigen (HBsAg) expressed by CHO cell lines. Also considering the effects of salt concentration and pH on HBsAg recovery and purification factor, orthogonal experiment design method was used to evaluated the absorbents. The results showed the butyl-S absorbent with the spacer arm length of C8, the ligand density of 22μmol/mL gel showed the best performance for the separation of HBsAg. Approximately 100% HBsAg recovery and 60 as purification fold were achieved by this media under the operating condition of pH 7.0 and 9% of salt concentrateion.