两种盖玻片对激光共聚焦显微镜成像效果的影响  

Effects of two different types of coverslip on immunofluorescene imaging with confocal microscopy

作　　者：骆宁[1] 范瑾瑾[1] 董秀清[1] 余学清[1] 

机构地区：[1]中山大学附属第一医院肾内科实验室,510080

出　　处：《新医学》2011年第7期433-435,F0003,共4页Journal of New Medicine

基　　金：国家985工程重点学科建设基金(2050205-203)

摘　　要：目的:探讨两种盖玻片制备的细胞爬片对激光共聚焦扫描显微镜(Confocal laserscanning microscopy,CLSM)采集免疫荧光图像效果的影响。方法:以适用于CLSM的专用盖玻片(A片)和组织病理学通用的普通盖玻片(B片)制备肾小管上皮细胞NRK52E的细胞爬片,分别进行胞浆蛋白转录激活因子3(STAT3),胞膜蛋白上皮型钙黏附蛋白(E-cadherin)及细胞骨架蛋白纤维状肌动蛋白(F-actin)的免疫荧光染色,在CLSM下以相同设置采集荧光图像并观察结果的异同。结果:CLSM图像显示,A片中胞浆蛋白STAT3弥散分布于胞浆,胞膜蛋白E-cadherin沿细胞连接处连续线状分布,细胞骨架蛋白F-actin向细胞表面伸出毛刺状突起的丝状伪足,均真实反映了蛋白的全部分布;而B片中胞浆蛋白STAT3弥散分布于胞浆,胞膜蛋白E-cadherin在细胞连接处呈不连续的点、短线状分布,细胞骨架蛋白F-actin未显示向细胞表面伸出的丝状伪足,不能完全真实反映蛋白的全部分布。结论:普通组织病理盖玻片用于CLSM的荧光图像观察,有时会使研究工作者在分析荧光图像时得出错误的结论,因此要选择适合CLSM的免疫荧光专用盖玻片,使实验条件趋于标准化。Objective ： To investigate the effects of two different types of coverslips on the immunofluorescence imaging with confocal microscopy. Methods： Both Coverslip A, which was the specified one for confocal microscopy, and Coverslip B, which was the common one used in routine histopathology, were applied for construction of cell-seeded coverslip for NRK52E cells. Protein STAT3, E-eadherin and F-actin, which located in cell cytoplasm, membrane and cytoskeleton, respectively, were detected with immunofluorescence staining. The images were collected by confocal microscopy with the same settings. Results： There was no difference in immunofluorescence pattern of cytoplastic protein STAT3 between images from Coverslip A and Coverslip B. Continuous linear distribution of E-cadherin on cytomembrane was observed in images from Coverslip, while dot-like or discontinuous linear pattern of E-cadherin was observed in images from Coverslip B. Filopodia like protrusions of F-actin on the cell surface were observed in images from Coverslip A, while no filopodia of F-aetin was observed in images from Coverslip B. Conclusion： Specialized eoverslip for confocal microscopy is necessary in immunofluorescence imaging in preventing misieading results.

关 键 词：盖玻片 激光共聚焦显微镜 免疫荧光 

分 类 号：R392.1[医药卫生—免疫学]