细胞的原子力显微镜最佳成像条件研究

The Research on best imaging Condition of the AFM on Cells

通过对不同处理条件和测试条件下人肝癌细胞SMMC-7721的原子力显微镜图像的分析和研究,得到了在大气环境和溶液环境中肝癌SMMC-7721细胞的最佳成像条件,同时建立了用原子力显微镜观测活细胞的实验方法.使用0.5%、1%、1.5%的戊二醛溶液固定细胞后再漂洗,变换原子力显微镜的扫描模式,调节扫描参量并在大气环境下观测以寻找该环境下的最佳成像条件;将用多聚赖氨酸处理基底后的培养细胞直接放置于生理溶液中用原子力显微镜进行溶液环境观测,比较扫描时限并分别改变环境液体类型、探针以及扫描频率,比较了不同条件下原子力显微镜图像的差异,在得出最优参量的同时对其相关原理进行了分析,建立了活细胞实时观测的实验方法.比较两种条件下的细胞图像发现,戊二醛溶液固定过的细胞与生理溶液环境中的活细胞有很大差别,在生理溶液条件下的细胞饱满,可见到光滑清晰的细胞边缘;但戊二醛溶液固定的细胞表面粗糙,细胞边缘不清晰,表明固定后观测到的细胞与生理状态下活细胞的表面形貌存在很大差异.

Through comparing the atomic force microscope （AFM） images of the 7721 cells under different treatment conditions and observation conditions,the best imaging condition of SMMC-7721 cells in the air and buffer solution was gotten, at the same time the experimental method of imaging living cells was established. Cells were fixed by 0.5 %, 1 %, 1.5 % glutaric dialdehyde, then washed by PBS three times, and observed in air circumstance with different scanning modes of AFM and scanning parameters to find the best figuring conditions in this circumstance. The living cells were cultured on the glass covered by polylysine, then put in the physiological solution and observed by the AFM directly. Compared the scanning time limit with changed the type of the circumstance solution, scanning tip and scanning frequency to get the optimized parameter. Images of the fixed cells were compared with the ones of living cells, and found very different. The living cells in the physiological solution were satiety and the edge was slick, but the edge and the surface of the fixed cells were rough and looked as fragmented.