论文部分内容阅读
目的 表达并纯化 9.1C3/LAIR 1胞膜外区与人的IgG1Fc段的融合蛋白 ,制备针对 9.1C3/LAIR 1胞膜外区的单克隆抗体 (mAb)。方法 构建真核表达载体 pIg/ 3C 9.1C3/LAIR 1,表达并纯化 9.1C3/LAIR 1 Fc融合蛋白。以 9.1C3/LAIR 1 Fc融合蛋白免疫BALB/c小鼠 ,制备。以间接免疫荧光染色和流式细胞术鉴定mAb的特异性 ,以竞争结合实验鉴定mAb识别LAIR 1的表位。结果表达并纯化了 9.1C3/LAIR 1 hIgFc融合蛋白 ,以其免疫BALB/c小鼠 ,获得 1株针对 9.1C3/LAIR 1胞膜外区的mAb 4H7。4H7对HL 6 0细胞和经 9.1C3/LAIR 1cDNA转染的COS7细胞上的表位识别 ,与已报道的抗 9.1C3mAb体不同。结论 成功地构建了 9.1C3/LAIR 1胞膜外区基因的真核表达载体 ,并表达纯化了 9.1C3/LAIR 1 hIgFc融合蛋白。获得一株针对 9.1C3/LAIR 1胞膜外区的mAb ,为进一步研究9.1C3/LAIR 1分子的结构和功能提供了新的手段
Objective To express and purify the fusion protein of 9.1C3 / LAIR1 extracellular domain and human IgG1Fc, and to prepare a monoclonal antibody (mAb) against the extracellular domain of 9.1C3 / LAIR1. Methods The eukaryotic expression vector pIg / 3C 9.1C3 / LAIR 1 was constructed and the 9.1C3 / LAIR 1 Fc fusion protein was expressed and purified. BALB / c mice were immunized with 9.1C3 / LAIR 1 Fc fusion protein. The specificity of mAb was identified by indirect immunofluorescence staining and flow cytometry. The binding of mAbs to LAIR 1 was identified by competitive binding assay. Results The 9.1C3 / LAIR 1 hIgFc fusion protein was expressed and purified, which was used to immunize BALB / c mice to obtain a mAb 4H7.4H7 against 9.1C3 / LAIR1 extracellular domain. / LAIR 1cDNA transfection of COS7 cells on the epitope recognition, and reported anti-9.1C3 mAb body different. Conclusion The eukaryotic expression vector of 9.1C3 / LAIR 1 extracellular domain gene was successfully constructed and the fusion protein of 9.1C3 / LAIR 1 hIgFc was expressed and purified. Obtaining a mAb against the extracellular domain of 9.1C3 / LAIR1 provides a new means for further investigation of the structure and function of the 9.1C3 / LAIR1 molecule