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目的:探讨胶质瘤细胞中p27kip1蛋白的表达、定位,为进一步研究p27kip1在胶质瘤发生、发展过程中的功能奠定理论基础。方法:用免疫荧光方法检测U87、LN308细胞p27kip1蛋白的定位情况;进一步分离两种细胞的细胞质与细胞核,在显微镜下观察细胞核形态并用DAPI染色分析细胞核完整性,提取蛋白用western blotting。方法检测分离的细胞质与细胞核蛋白的纯度,并检测p27kip1在细胞中的表达情况。结果:成功分离了细胞的细胞浆与细胞核,并得到纯度较好的细胞浆蛋白与细胞核蛋白。确定了p27kip1蛋白主要表达于U87和LN308细胞的细胞质中。结论:p27kip1蛋白在恶性胶质瘤中可能主要表达在细胞质中,并且其亚细胞定位可能与胶质瘤的恶性程度相关。
OBJECTIVE: To investigate the expression and localization of p27kip1 in glioma cells and to lay a theoretical foundation for further study on the function of p27kip1 in the development and progression of glioma. Methods: The localization of p27kip1 protein in U87 and LN308 cells was detected by immunofluorescence. The cytoplasm and nucleus of both cells were further separated. The morphology of nucleus was observed under microscope and the nuclear integrity was analyzed by DAPI staining. The protein was extracted by western blotting. Methods The purity of isolated cytoplasm and nucleoprotein was detected and the expression of p27kip1 in cells was detected. Results: The cytoplasm and nucleus of cells were successfully separated and the cytoplasm and nucleoprotein with good purity were obtained. It was confirmed that p27kip1 protein is mainly expressed in the cytoplasm of U87 and LN308 cells. Conclusion: p27kip1 protein may be mainly expressed in cytoplasm in malignant gliomas, and its subcellular localization may be related to the malignancy of glioma.