摘要
目的:建立一种基于免疫荧光图像的快速、简便、半定量分析微管蛋白的方法.方法:小鼠成骨样细胞MC3T3分别在正常及三维回转条件培养48h后,免疫荧光细胞化学技术标记对照组和回转组的成骨细胞微管骨架,激光共聚焦显微镜采集荧光图像;采用三种不同的专业图像分析软件SimplePCI,Imagepro-Plus6.0(IPP)及ImageJ定量分析三维回转培养条件对小鼠成骨样细胞MC3T3-E1微管细胞骨架的影响;用Western Blot检测α-微管蛋白表达量;用ImageJ软件确定MC3T3-E1细胞微管骨架的分形维数.结果:对采集到的细胞骨架荧光染色图像做定量分析发现,与对照组相比,经过48h三维回转培养,MC3T3-E1细胞图像的荧光强度明显降低(P<0.01);Western Blot结果表明,经过48h三维回转培养,MC3T3-E1细胞α-微管蛋白表达量表达量明显下降;Im-ageJ软件分析结果表明,经过48h三维回转培养,MC3T3-E1细胞微管的分形维数也明显低于对照组(P<0.05).结论:图像分析软件的结果和实验结果一致,表明基于免疫荧光图像的分析方法是可行的,为细胞形态学的定量研究提供了新思路和手段.
AIM：To establish a quick,convenient and semi-quantitative method based on fluorescent images for microtubule cytoskeleton analysis.METHODS：Three image analysis software including SimplePCI,Image J and Image-pro plus 6.0（IPP）,and Western Blot assays were used to qualify the microtubule cytoskeleton alterations in MC3T3-E1 cell after 48 h of being cultured under Radom Position Machine（3D） condition.The Fractal dimensions（D） of cytoskeleton architecture of MC3T3-E1 were calculated by box counting method using Image J.RESULTS：After being cultured for 48 h in RPM,the Fluorescent intensity（grey level） of MC3T3-E1 cytoskeleton was lower than that under control condition（P0.01）.Western Blot analysis showed after being cultured for 48 h in RPM,α-tubulin in MC3T3-E1 cells expression was decreased significantly（P0.001）,and the fractal dimension was also lower than that under control condition（P0.05）.CONCLUSION：The method based on fluorescent Images used in this study is useful to qualify both the location and expression of microtubule cytoskeleton,which provides a new insight into the quantitative morphological study of cytoskeleton.
出处
《第四军医大学学报》
北大核心
2009年第24期2901-2904,共4页
Journal of the Fourth Military Medical University
基金
国家自然科学基金面上项目(30840030)
863重大项目子课题(2008AA12A218)
关键词
荧光图像
定量
微管
细胞骨架
分形维数
fluorescent image
semi-qualification
microtubule
cytoskeleton
fractal dimensions
