Calcein-AM和Annexin V在流式细胞术检测早期细胞凋亡的敏感性比较

Comparison of Calcein-AM and Annexin-V as early vital markers of apoptosis by cytometry

目的比较Calcein-AM与Annexin V用于流式细胞术检测活细胞早期凋亡时的敏感性,建立新的检测细胞早期凋亡的方法。方法将人类急性T淋巴细胞白血病细胞株(Molt-4)经喜树碱(CPT)、紫外线(UV)诱导后,分别采用Calcein-AM及异硫氰酸荧光素标记的膜联蛋白V(Annexin V)染色,然后用流式细胞仪分析凋亡率,通过比较诱导后不同时间点的凋亡率来分析Calcein-AM和Annexin V对于检测细胞早期凋亡的敏感性。结果在喜树碱诱导模型中,加药后2 h即可用Calcein-AM检测到明显的细胞凋亡,而3 h后才能用Annexin V检测到;在紫外线诱导模型中,照射后1 h即可用Calcein-AM检测到明显的细胞凋亡,而要在2 h后才能用Annexin V检测到。结论Calcein-AM用于检测活细胞早期凋亡比Annexin V更为敏感,是一种稳定、可靠、灵敏度高并且较为经济的检测细胞早期凋亡的活细胞染料。

Objective The current study was designed to compare the sensitivities of Calcein-AM and Annexin V-FITC for early detectionof apoptosis in living cells by flow cytometry and to establish a new method for early detection of apoptosis. Methods The Molt-4 cellline treated with camptothecin （CPT） or ultraviolet（UV） were labeled with Calcein-AM and FITC-conjugated Annexin V respectively,then the apoptosis rate were analyzed by flow cytometry and the performance of Calcein-AM and Annexin V for early detection of apoptosis were compared. Results Our results showed that both probes allowed the detection of apoptotic cells. I^Iowever, Calcein-AM was moresensitive than Annexin V. We could detect the apoptosis induced by CPT in two hours after inducing with Calcein-AM and in threehours after inducing with Annexin V. The apoptosis induced by UV could be detected in one hours after inducing with Calcein-AM buttwo hours after inducing with Annexin V. Conclusion Annexin V is less sensitive than Calcein-AM for detection of early apoptosis, andthe new method for detection of early apoptosis with Calcein-Am is stable,reliable, sensitive and low-cost.