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目的 :建立稳定表达人巨噬细胞集落刺激因子受体 (M- CSFR)或人巨噬细胞集落刺激因子受体—膜结合型巨噬细胞集落刺激因子 (M- CSFR- m- M- CSF)的人—鼠嵌合肿瘤细胞系 ,作为研究肿瘤免疫治疗的实验模型。方法 :用 PCR技术构建 M- CSFR- m- M- CSF融合 c DNA的真核表达载体。将携带 M- CSFR及 M- CSFR- m- CSF的重组质粒转染小鼠骨髓瘤细胞系 SP2 / 0 ,G4 18筛选所得的混合克隆 ,有限稀释后克隆化培养。结果 :得到 3株稳定表达人 M- CSFR和 4株稳定表达人 M- CSFR- m- M-CSF的嵌合肿瘤细胞系。ABC免疫酶标法和 RT- PCR法证实所获细胞系有明显的目的蛋白表达。皮下接种 BAL B/ c小鼠成瘤率 10 0 %。结论 :建立了稳定表达人 M- CSFR或人 M- CSFR- m- M- CSF的人—鼠嵌合肿瘤细胞系 ,并已成功地用于 M- CSF及其受体 DNA疫苗免疫效应的研究。其原理和方法可供同类研究工作借鉴。
OBJECTIVE: To establish a cell line stably expressing human macrophage colony-stimulating factor receptor (M-CSFR) or human macrophage colony-stimulating factor receptor-membrane-bound macrophage colony stimulating factor (M-CSFR-m- Human-mouse chimeric tumor cell lines as an experimental model for the study of tumor immunotherapy. Methods: The eukaryotic expression vector of M-CSFR-m-M-CSF fusion cDNA was constructed by PCR. The recombinant clones carrying M-CSFR and M-CSFR-m-CSF were transfected into mouse myeloma cell lines SP2 / 0 and G418. The resulting mixed clones were cloned and subcultured in a limited dilution. Results: Three chimeric tumor cell lines stably expressing human M-CSFR and four stably expressing human M-CSFR-m-M-CSF were obtained. ABC immunoenzyme method and RT-PCR method confirmed that the cell lines obtained had a significant expression of the target protein. BALB / c mice were inoculated subcutaneously with a tumorigenic rate of 10%. Conclusion: A human-mouse chimeric tumor cell line stably expressing human M-CSFR or human M-CSFR-m-M-CSF has been established and has been successfully used in the study of the immune effects of M-CSF and its receptor DNA vaccines . Its principles and methods can be used for reference for similar research.