【摘要】 为找出准确定量分析细胞脂滴的方法,以猪卵母细胞为模型,利用2种非破坏性的形态学方法——三维重建分析法和平面光密度法,通过荧光染料特异性标记脂滴,对单个细胞内脂滴含量进行准确定量和对比分析。结果表明:2种方法都能根据荧光图像定量细胞内脂滴,并统计出脂滴大小、密度和分布规律等指标;随机选取100个脂滴含量不等的卵母细胞,三维重建分析法能更准确地反映样本脂滴含量变异情况(变异系数C.V.=0.86),而光密度法的C.V.=0.68;针对同一细胞不同角度的重复观察,三维重建分析法具有更好的重复性(变异系数C.V.=0.07,组内相关系数ICC>0.9),而光密度法C.V.=0.37、ICC<0.4。因此基于激光共聚焦扫描的三维重建法能够更准确地对细胞内的脂滴含量进行定量分析。更多还原

【Abstract】 Lipid droplets are considered to play important roles in cells as a dynamic organelle,therefore the quantification of intracellular lipid droplets is significantly important.In this research,pig oocytes were chosen as a model,while the lipid drop was specially marked by the fluorescent dye.Two methods of 3Dreconstruction method and the flat optical density method were conducted to overcome the challenges of destroying the structure of the cell for lipid drop quantitative and comparative analysis in a single cell.The results showed that both of the methods could quantify intracellular lipid droplets according to the fluorescence image,providing the statistics of lipid droplet size,density,distribution of index.Randomly selecting 100 oocytes which had different contents of lipid droplets,3Dreconstruction method could reflect the variation of the content of lipid droplets more accurately(coefficient of variation C.V.=0.86),but the flat optical density method C.V.=0.68.Scanning the same cell in multiple angles for repeat observation,3D reconstruction method had higher repeatability(coefficient of variation C.V.= 0.07,interclass correlation coefficient ICC>0.9),but the flat optical density method C.V.=0.37,ICC<0.4.The method of 3Dreconstruction based on the confocal laser scanning can quantify the content of lipid droplets in the cell with higher accuracy and reproducibility.更多还原