中文摘要：

目的探索建立人CD123稳定表达细胞株的方法,为进一步实现人CD123的抗体规模化生产提供材料。方法构建人CD123 CDS区真核表达载体（hCD123 CDS-pEGFP-N1）;经脂质体包裹转染中国仓鼠卵巢细胞CHO-S;经流式分选结合G418筛选,并用流式细胞术和Western印迹法检测人CD123蛋白的表达情况,初步筛选出人CD123高表达的CHO-S细胞株;并进一步用经人CD123 Fc重组融合蛋白免疫的小鼠血清检测该细胞株hCD123表达情况。结果测序及GenBank中序列比较结果显示扩增到的人CD123基因序列是正确的,酶切结果表明表达质粒构建正确;流式分选结合G418筛选得到1个人CD123高表达的细胞株——克隆8G9 CHO-S,其CD123-APC阳性细胞百分数为92.07%,用融合蛋白免疫的血清检测各细胞株人CD123表达情况,结果显示克隆8G9 CHO-S细胞和对照CHO-S细胞的CD123阳性率分别为（97.94±1.93）%和（2.84±1.35）%,差异具有统计学意义（P〈0.01）。结论建立了人CD123抗原稳定高表达的细胞株克隆8G9 CHO-S,为进一步实现人CD123的抗体规模化生产及其导向治疗奠定了坚实的基础。

英文摘要：

Objective To establish a cell line that can stable express hCD123,which can be applied in the large-scale production of anti-CD123 antibody.Methods Expression vector of hCD123 CDS-pEGFP-N1 was constructed and transfected into chinese hamster ovarian cell line CHO-S using Lipofectamine 2000.G418 was used to select cell clones that could stably express human CD123.The expression of CD123 was verified by Western blot and flow cytometry.The cells that could highly express human CD123 were isolated by Flow cytometry and successfully established as CD123＋ CHO-S cell line.Furthermore,the sera of the mice immunized with recombinant fusion protein hCD123-Fc were employed to detect the expression of hCD123 protein in such CHO-S cell line.Results The sequence of human CD123 in the experiment was identical to the sequence in the GeneBank,and restriction enzyme digestion results indicated successful construction of CD123 expression vector.A stable cell line clone 8G9 CHO-S that highly expressed human CD123 was obtained by G418 positive selection and flow cytometry.CD123 positive ratio was 92.07%.Using the serums of the mice immunized with recombinant fusion protein hCD123-Fc to analysis the expression of hCD123 protein in various cell lines,the data showed there was around（97.94±1.93）% CD123＋ cells in 8G9 CHO-S cell line compared with a（2.84±1.35）% positive rate in control CHO-S cells（P〈0.01）.Conclusion A human cell line 8G9 CHO-S that can stable express CD123 was established,providing a significant prerequisite in the future large-scale production of anti-CD123 antibody and the related targeted therapy