中文摘要：

目的探讨在激光扫描共焦显微镜（LSCM）观测活细胞的过程中,激光对细胞内反应氧（ROS）、细胞内钙离子浓度（[Ca^2＋]i）和细胞膜完整性的影响。方法用H2DCFDA、Fluo-4AM和Calcein-AM/PI分别检测488nm激光对脐静脉内皮细胞株（ECV-304）细胞内ROS、[Ca^2＋]i和细胞膜完整性的影响,Ultra view LCI（live cell image）共聚焦成像系统采集并分析结果。结果激光照射后细胞内DCF的荧光强度迅速上升,约80s达到峰值,随后下降并逐渐回到基础水平;Fluo-4的荧光强度在70min内持续平缓地上升;Calcein的荧光强度明显下降,少量细胞PI阳性染色。结论488nm激光照射可引起细胞内ROS和[Ca^2＋]i的增加,但对细胞膜的完整性没有显著影响。

英文摘要：

Objective To investigate the effects of laser irradiation on intracellular ROS （reactive oxidant species ）, intracellular calcium concentration（ [ Ca^2＋ ]i, and cell membrane integrity in the process of live cell imaging with confocal laser scanning microscopy. Methods The effects of a given laser irradiation on ROS, intraeellular calcium concentration （ [ Ca^2＋ ] i and cell viability were revealed respectively by stained ECV-304 with Hz DCFDA, Fluo-4AM and calcein-AM/PI, and visualized and analyzed using ultra view LCI（live cell image）confocal microscopy. Results The irradiation of 488nm laser induced fluorescent intensity of DCF to increase abruptly and attain the climax in about 80 seconds, afterwards the fluorescent intensity fell and returned to the baseline. In the 70 minutes of the irradiation, the fluorescent intensity of intracellular Fluo-4 kept a slightly ascending tendency. The fluorescent intensity of ealeein decreased 15minutes after the irradiation, and serval cells were PI positively stained. Conclusion 488nm laser irradiation induces intracellular reactive oxidant species （ROS） and calcium concentration to increase, but there is no significant influence on cell membrane integrity.