细胞骨架染色中不同固定方法的探讨  

A Study of Different Fixatives in Cytoskeleton Staining

作　　者：张维[1] 骞爱荣[1] 高翔[1] 胡丽芳[1] 瓮媛媛[1] 韩晶[1] 商澎[1] 

机构地区：[1]西北工业大学生命科学院,西北工业大学特殊环境生物物理学研究所,空间生物实验模拟技术国防重点学科实验室,西安710072

出　　处：《中国细胞生物学学报》2010年第1期149-153,共5页Chinese Journal of Cell Biology

基　　金：国家高技术研究发展计划(863计划)(No.2008AA12A218);国家自然科学基金(No.30840030)资助项目~~

摘　　要：探讨不同细胞固定方法对细胞骨架染色的影响,以期发现适合于微丝与微管细胞骨架染色的固定方法。本文分别采用0.5%戊二醛、95%乙醇、4%甲醛及无水冷丙酮等四种固定液对人骨肉瘤细胞MG-63进行固定,然后分别对微丝和微管进行染色、封片,采用激光共聚焦扫描显微镜观察细胞微丝及微管骨架的形态。同时采用图像分析方法对细胞骨架微丝、微管荧光图像进行定量分析。结果表明,0.5%戊二醛固定的细胞,微丝呈清晰的束状纤维,微管呈有规律的束状纤维;95%乙醇固定的细胞,微丝纤维连贯性差,微管纤维非常清晰;4%甲醛固定的细胞,微丝纤维之间比较模糊,微管纤维呈圆弧状,联贯性较差;无水冷丙酮固定的细胞,微丝明显皱缩,呈模糊状,微管清晰可见,连贯性较好。因此,不同固定液固定的原理不同,要根据实验目的,选择合适的固定方法。The purpose of this study is to discuss the effects of different fixatives on cytoskeleton staining and to find out the suitable fixative methods of F-actin and microtubule staining. Four fixatives, 0.5% glutaraldehyde, 95% alcohol, 4% formaldehyde and acetone, were used to fix MG-63 cells respectively, and then F-actin and microtubule were stained, F-actin and microtubule images captured by confocal laser scanning microscopy were observed and quantitatively analyzed. The results showed that F-actin filaments and microtubules of the MG-63 cells fixed by 0.5% glutaraldehyde were bunchy. The coherence of F-actin filaments of the MG-63 cells fixed by 95% alcohol was poor but microtubules were very distinct. F-actin filaments of the cells fixed by 4.% formaldehyde were blurred, and microtubules were around the nucleus and became discontinuous. F-actin filaments of the cells fixed by acetone were dramatically disrupted but microtubules were very clear. The principle of the different fixative was different, so it is necessary to select the suitable fixative method according to the purpose of the experiment.

关 键 词：细胞骨架 固定方法 激光共聚焦显微术 人骨肉瘤细胞 

分 类 号：Q245[生物学—细胞生物学]