卵巢癌单克隆抗体杂交瘤细胞183B2的无血清培养研究

Serum-free culture of hybridoma cell line 183B2 with ovarian carcinoma monoclonal antibody

目的 建立体外无血清培养杂交瘤细胞生产单克隆抗体的方法 ,并进行纯化工艺的探讨 ,为规模化抗体生产奠定基础。方法 以卵巢癌单克隆抗体杂交瘤细胞 183B2为研究对象 ,通过与常规的有血清培养法比较 ,研究了无血清培养基H -SFM中杂交瘤细胞的生长特性、抗体分泌效率、活性 ,用Supper-Spinner搅拌瓶进行了放大培养并用ProteinA亲和层析法对无血清培养上清中抗体的纯化工艺进行了探索。结果 183B2能够直接适应无血清培养基 ,不需要驯化 ;抗体生成约 4d达到高峰 ,抗体产量可达 5 0mg/L/ 10 6个细胞 ,无血清培养基中细胞生长及单抗的产量与效价与有血清培养的相比没有明显差别。用 1LSupper-Spinner搅拌瓶无血清放大培养 183B2细胞成功 ,培养上清经ProteinA亲和层析纯化 ,蛋白产率约 10mg/L培养上清。结论 本研究初步成功探索了卵巢癌单克隆抗体细胞 183B2的无血清培养方法和抗体提纯工艺 ,为进一步大规模抗体生产提供了实验依据。

Objective To culture the hybridoma cells in serum-free medium (SFM) and to purify the monoclonal antibody against ovarian carcinoma from the culture supernatant in vitro.Methods Ovarian carcinoma hybridoma cell 183B2 was cultured in Hybridoma-SFM (H-SFM), and the control was cultured with RPMI1640 medium supplemented with 10% fetal bovine serum. The characters of cell growth and antibody production were compared. The protein was purified from the culture supernatant with Protein A affinity chromatography.Results Ovarian carcinoma hybridoma cell 183B2 was successfully adapted to grow in H-SFM. Cell growth rate, monoclonal antibody (Mab) production and Mab activity were similar between the H-SFM and the control. The peak of IgG, which was approximately 50 mg.L -1 /10 6 cells, was obtained after 4 days culture. About 10 mg.L -1 (cell culture supernatant) of the Mab could be achieved.Conclusions The 183B2 hybridoma cells can be cultured in H-SFM and the Protein A chromatography can be applied to purify the Mab from the cell culture supernatant. These results obtained in this study offer practical methods for scale-up Mab production.