Photochemically generated cytosolic calcium pulses and their detection by fluo-3.
Photochemically generated cytosolic calcium pulses and their detection by fluo-3.
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DOI：
10.1016/s0021-9258(18)83166-0
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发表时间：
1989-05
期刊：
影响因子：
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通讯作者：
J P Y Kao;A. Harootunian;R. Tsien
中科院分区：
文献类型：
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作者：
J P Y Kao;A. Harootunian;R. Tsien
Fluo-3，在可见光波长下可激发的新型荧光Ca 2+指示剂家族的一个成员（Minta，A.，Kao，J.P.Y.，和Tsien，R. Y.（1989）J.Biol.Chem.264，8171-8178），已经在活细胞中进行了测试。我们证明，荧光-3可以加载到成纤维细胞和淋巴细胞与pentaacetoxymethyl酯的染料孵育，酯水解胞内产生真正的荧光-3能够指示的变化[Ca 2 +] i诱导的激动剂刺激。Fluo-3也可以与光不稳定化合物如nitr-5和笼状三磷酸肌醇一起沿着显微注射到成纤维细胞中用于光释放实验。Fluo-3允许连续监测[Ca 2 +] i，而不干扰使用UV敏感的笼状化合物。还描述了结合使用离子载体和重金属离子在实验结束时校准fluo-3强度以得到[Ca 2 +] i的程序。
Fluo-3, one member of a family of new fluorescent Ca2+indicators excitable at wavelengths in the visible (Minta, A., Kao, J. P. Y., and Tsien, R. Y. (1989)J. Biol. Chem.264, 8171–8178), has been tested in living cells. We demonstrate that fluo-3 can be loaded into fibroblasts and lymphocytes by incubation with the pentaacetoxymethyl ester of the dye and that the ester is hydrolyzed intracellularly to yield genuine fluo-3 capable of indicating changes in [Ca2+]iinduced by agonist stimulation. Fluo-3 can also be microinjected into fibroblasts along with photolabile compounds such as nitr-5 and caged inositol trisphosphate for photorelease experiments. Fluo-3 permits continuous monitoring of [Ca2+]iwithout interference with use of UV-sensitive caged compounds. A procedure for combined use of ionophore and heavy metal ions in end-of-experiment calibration of fluo-3 intensities to give [Ca2+]iis also described.