摘要
目的构建适于普通小鼠的癌胚抗原（CEA）阳性结肠癌细胞株。方法以重组CEACAM5cDNA-Lentivims慢病毒转染小鼠结肠癌CT26细胞．抗生素筛选2周．以有限稀释法获得癌胚抗原阳性细胞CT26CEA，体外培养至第7代和第14代进行鉴定。用RT．PCR法和Western blot法分别检测CEACAM5mRNA及蛋白表达．用荧光显微镜和免疫细胞化学法观察CEACAM5表达位置：并以第14代CT26CEA细胞建立BALB／c鼠皮下种植瘤模型．用活体荧光成像法和免疫组织化学染色观察瘤体CEACAM5表达情况。结果在第7代和第14代CT26CEA细胞内可以检测到CEACAM5mRNA及CEACAM5蛋白表达．荧光显微镜和免疫细胞化学法观察到其表达主要在细胞质，活体荧光成像法和免疫组织化学染色观察到小鼠瘤体有丰富CEACAM5表达。结论成功构建小鼠结肠癌CT26CEA细胞株，能够在体外培养及小鼠体内稳定表达癌胚抗原．为研究正常免疫环境下癌胚抗原对结肠癌进展的影响及作用机制提供了适用工具．
Objective To establish a colon cancer cell line with stable expression of carcinoembryonic antigen （CEA）. Methods Recombinant lentivirus conjugated with CEACAM5 cDNA were used to transfect wild CT26 ceils. Antibiotics were given for 2 weeks to select CEA positive cells. A single transfected clone was obtained using limiting dilution. The 7th and 14th passages of cells cultured in vitro were detected for CEACAM5 mRNA by RT-PCR, and protein by westem blot. The location of CEACAM5 expression was examined using fluorescent microscope and immunocytochemistry. The 14th passage cells were injected subcutaneously into mice to create BALB/c model and CEACAM5 protein was detected by in vivo fluorescence image analysis system and immunohistoehemistry： Results CEACAM5 mRNA and protein were found in the 7th and 14th passages of CT26CEA cells, which were proved to locate in the cytoplasm by fluorescence microscope and immunohistoehemistry. Abundant CEACAM5 protein was found in subcutaneous tumors by in vivo fluorescence image analysis system and immunohistochemistry. Conclusion Colon cancer cell line CT26 with stable expression of CEA in vitro and in mice can be used as a suitable tool to facilitate research on the impact and mechanism of CEA on colon cancer under normal immune environment.
出处
《中华胃肠外科杂志》
CAS
北大核心
2011年第5期360-363,共4页
Chinese Journal of Gastrointestinal Surgery
基金
国家自然科学基金资助（30560150）
关键词
结肠肿瘤
癌胚抗原
CT26细胞
模型
小鼠
Colonic neoplasms
Carcinoembryonie antigen
CT26 cells
Models, mouse
