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目的探讨CHO-HB2细胞株表达抗HBsAg人IgG的影响因素。方法观察CHO-HB2细胞表达抗体的稳定性不同培养基以及添加剂的效用细胞接种密度和培养持续时间的选择金属离子Zn2+诱导表达的作用,以及滚瓶的培养效果。结果CHO-HB2细胞的亚克隆株F4经液氮冻存半年多复苏后,在无氨甲喋呤MTX的培养基中连续传代第2个月,抗体表达量开始下降,5个月时接近用MTX施加选择压力前的基础水平。在培养基中添加多种氨基酸和维生素,并加入适量的Zn2+诱导金属硫蛋白启动子,可使表达量明显增加。滚瓶培养可进一步增加抗体的表达量,在无蛋白的无血清培养基中的表达量,从6.2mg/L提高到28mg/L。结论应及时用MTX对细胞株进行再加压筛选,以保持细胞稳定表达抗体的能力。通过优化培养条件,使用完全无蛋白的无血清培养基,可收到较好的效果,且有利于表达抗体的纯化。
Objective To investigate the factors affecting the expression of anti-HBsAg human IgG in CHO-HB2 cell line. Methods The stability of antibody expressed in CHO-HB2 cells was observed. 效 The effects of different media and additives 选 The selection of cell seeding density and culture duration The role of Zn2 + -induced expression of metal ions and the culture effect of roller bottles. Results The subclone F4 of CHO-HB2 cells was sub-cloned in liquid nitrogen for more than six months and then resuspended for more than six months. The cells were continuously passaged in the medium without methotrexate (MTX) for 2 months, and the antibody expression began to decrease. At 5 months, MTX choose to exert pressure before the basic level. In the medium to add a variety of amino acids and vitamins, and add the appropriate amount of Zn2 + -induced metallothionein promoter, can make a significant increase in expression. The roller bottle culture further increased the expression level of the antibody, and the expression level in the protein-free serum-free medium increased from 6.2 mg / L to 28 mg / L. Conclusion MTX cell lines should be repressurized in time to maintain the ability of cells to stably express the antibody. By optimizing the culture conditions, the use of completely protein-free serum-free medium, can receive better results, and is conducive to the expression of antibody purification.