应用激光扫描共聚焦显微镜观察晶体上皮细胞听游离钙

用钙的荧光批示剂ｆｌｕｏ－３和激光扫描共聚焦显微镜观察大白鼠的晶体上皮细胞中的游离钙。方法 ｆｌｕｏ－３着染细胞后，应用ＬＳＣＭ观察细胞内ｆｌｕｏ－３在钙螯合后的荧光分布，最后用钙离子载体Ａ２３１８７和重金属离子Ｍｎ＾＋＋作校正，将ｆｌｕｏ－３与钙结合显示的荧光强度转换为「（Ｃａ＾＋＋）」值。

Observation of the free calcium of lens epithelial cell using laser scanning confocal microscopy]

OBJECTIVE: Using a new fluorescent Ca++ indicator (fluo-3) and laser scanning confocal microscopy (LSCM) to monitor the intracellular Ca++ of rat lens epithelial cell (LEC). METHODS: The LEC was loaded with fluo-3, then the distribution of free Ca++ in the LEC was observed by using LSCM monitoring the distribution of fluorescence that is given due to the chelation between fluo-3 and Ca++. At last, ionophore (A23187) and heavy-metal ions (Mn++) were applied to calibrate fluo-3 intensities to obtain Ca++]i value. RESULTS: The free calcium in LEC is mainly located in the nucleus. Free calcium concentration of rat LEC is 259.79 +/- 49.24 nmol/L. CONCLUSION: Fluo-3 can be loaded into LEC by incubation with acetoxymethyl (AM) ester of the dye, and LSCM can be used to observe the cellular free calcium directly by surveying the fluorescence. This method will pave a way technically for the further study of intracellular free Ca++ of the LEC.