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目的 在 2 93细胞中表达人源化单链抗体 /人白细胞介素 2 (hIL 2 )双功能融合蛋白及评价其生物学活性。方法 将构建的表达载体转染 2 93细胞 ,G418筛选阳性克隆建立稳定表达细胞系 ,免疫印迹法 (Westernblot)、酶联免疫吸附试验 (ELISA )及 [3 H]脱氧胸苷 (3 H TdR)渗入法检测细胞上清中融合蛋白浓度和生物学活性。结果 融合蛋白浓度达 (10 2 .0± 4.2 ) g/L ,5 μl细胞上清即可在 45× 10 3 处呈现清晰蛋白条带。其中hIL 2无活性丢失 ,与标准对照比较差异无显著性 (P >0 .0 5 ) ;抗体的抗原结合活性有部分下降 ,与标准对照比较差异有非常显著性 (P <0 .0 1)。结论 该融合蛋白可在 2 93细胞系中得到高效表达 ,其中hIL 2部分无活性丢失 ,抗体的活性有部分下降
Objective To express human scFv / human interleukin 2 (hIL 2) bifunctional fusion protein in 293 cells and evaluate its biological activity. Methods The constructed expression vector was transfected into 293 cells, and the positive clones were screened by G418 to establish stable cell lines. Western blotting, ELISA and [3H] - deoxythymidine (3 H TdR) Infiltration method was used to detect the concentration and biological activity of the fusion protein in the cell supernatant. Results The fusion protein concentration was (10 ± 0.2 ± 4.2) g / L, and 5 μl of cell supernatant showed a clear protein band at 45 × 10 3. The activity of hIL 2 was lost, with no significant difference from the standard control (P> 0.05). The antigen binding activity of the antibody partially decreased, with significant difference compared with the standard control (P <0.01) . Conclusion The fusion protein can be highly expressed in 293 cell line, in which the hIL 2 part is inactive and the antibody activity is partially decreased