优化大鼠肺动脉平滑肌细胞骨架F-actin图像的激光共聚焦显微技术

摘要

Objective: To optimize the techniques of laser confocal scanning microscope (LCSM) on the image of F-actin in PASMCs. Method: Using different concentrations of fluorescent dyes and different incubation conditions, the image of F-actin in PASMCs was observed with CLSM. Results: Final concentration of 10 μg/mL FITC-Phalloidin, 10 μg/mL PI, stained for 30 minutes, and worked by the Rnase at 37 ℃ were the optimal experimental conditions. The image showed that F-actin was filamentous green fluorescent light, PI labeled nuclei red fluorescence, and no significant PI staining cytoplasm. Conclusion: Through the final optimization of dyeing conditions, the experiment showed the superiority of the techniques of LCSM to observe F-actin in plane and three-dimensional cytoskeleton, which provided a good technical platform for further experiments.%目的:优化大鼠肺动脉平滑肌细胞(PASMCs)骨架蛋白F-actin图像的激光共聚焦显微技术.方法:采用不同浓度梯度的荧光染料及不同孵育条件,应用激光共聚焦显微扫描技术对PASMCs细胞骨架F-actin进行形态学现察.结果:终浓度10 mg/mL FITC-鬼笔环肽及10 mg/mL PI,分别染色30 min,并经RNA酶37 ℃处理为最适实验条件,可见F-actin呈明亮细丝状绿色荧光,PI标记的胞核呈红色荧光,胞浆无明显PI染色.结论:通过对染色条件的最终优化,表现出激光共聚焦技术在平面及三维同时现察细胞骨架F-actin的优越性,为进一步实验提供良好的技术平台.