Comparison of annexin V and Calcein-AM as early vital markers of apoptosis in adherent cells by confocal laser microscopy
Comparison of annexin V and Calcein-AM as early vital markers of apoptosis in adherent cells by confocal laser microscopy
复制标题
DOI：
10.1177/002215549804600804
复制
发表时间：
1998-08-01
影响因子：
3.2
通讯作者：
Gazzola, GC
中科院分区：
文献类型：
--
作者：
Gatti, R;Belletti, S;Gazzola, GC
虽然细胞凋亡的形态学标准一般是可靠的，迄今为止尚未进行系统的比较所采用的技术。在这项研究中，使用共聚焦激光显微镜，我们比较了膜联蛋白V-FITC和钙黄绿素-AM的性能，在活的贴壁细胞凋亡的早期检测。实验在两种不同的细胞系PC 12和NIH 3 T3上进行，这两种细胞系具有不同的形状和粘附特性。凋亡的过程中，随后在一个预定的领域的相同的细胞通过一个特殊的流动室的延长的时间。我们的研究结果表明，这两种探针允许在任一细胞系中检测凋亡细胞。然而，在钙黄绿素可视化上清楚地表现出凋亡变化的一些细胞是膜联蛋白阴性的。在NIH 3 T3细胞中，凋亡细胞的膜联蛋白阴性与细胞形状和粘附特性的保持相关。这些发现表明，至少在PC 12和NIH 3 T3细胞中，膜联蛋白可能比钙黄绿素-AM对早期凋亡检测的敏感性低，并且对于NIH 3 T3细胞，表明磷脂酰丝氨酸暴露在某种程度上与细胞形状和/或对培养基质的粘附的变化有关。
Although morphological criteria for apoptosis are in general reliable, no systematic comparison of the techniques employed thus far has yet been performed. In this study, using confocal laser microscopy, we compared the performance of annexin V-FITC and calcein-AM for early detection of apoptosis in living adherent cells. Experiments were carried out on two distinct cell lines, PC12 and NIH3T3, endowed with different shape and adhesion properties. The apoptotic process was followed for a prolonged period in the same cells of a predetermined field by means of a special flow chamber. Our results show that both probes allowed the detection of apoptotic cells in either cell line. However, some cells that clearly exhibited apoptotic changes on calcein visualization were annexin-negative. In NIH3T3 cells, annexin negativity of apoptotic cells was correlated with the preservation of cell shape and adhesion properties. These findings show that, at least in PC12 and NIH3T3 cells, annexin might be less sensitive than calcein-AM for early apoptosis detection and, for NIH3T3 cells, suggest that phosphatidilserine exposure is in some way linked to changes in cell shape and/or adhesion to culture substrate.