MEMBRANE-POTENTIAL CAN BE DETERMINED IN INDIVIDUAL CELLS FROM THE NERNSTIAN DISTRIBUTION OF CATIONIC DYES
MEMBRANE-POTENTIAL CAN BE DETERMINED IN INDIVIDUAL CELLS FROM THE NERNSTIAN DISTRIBUTION OF CATIONIC DYES
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DOI：
10.1016/s0006-3495(88)83158-8
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发表时间：
1988-05-01
影响因子：
3.4
通讯作者：
LOEW, LM
中科院分区：
文献类型：
--
作者：
EHRENBERG, B;MONTANA, V;LOEW, LM
选用阳离子荧光染料的分布可用于用微荧光计测量单个细胞的膜电位。这些染料的基本属性包括膜渗透性，低膜结合，对环境不敏感的光谱特性，当然还有强荧光。在海拉细胞上筛选了一系列染料，以满足这些标准，并发现几种市售染料是令人满意的。此外，还通过四甲基罗丹明的酯化反应合成了两种新的染料。对所测荧光强度的分析需要对从细胞聚焦平面外收集的荧光和染料的非电位结合进行校正。测量和分析是在三种不同的细胞类型上进行的，其中存在大量关于膜电位的文献；这部作品所确定的潜力始终在文学价值的范围内。罗丹明酯是一种无毒的高荧光染料，不会形成聚集体，也不会在荧光效率上显示依赖于结合的变化。因此，它们的可逆积累与细胞内和细胞外荧光的对比定量相关，并允许连续监测单个细胞的膜电位。
The distribution of a selection of cationic fluorescent dyes can be used to measure the membrane potential of individual cells with a microfluorometer. The essential attributes of these dyes include membrane permeability, low membrane binding, spectral properties which are insensitive to environment, and, of course, strong fluorescence. A series of dyes were screened on HeLa cells for their ability to meet these criteria and several commercially available dyes were found to be satisfactory. In addition, two new dyes were synthesized for this work by esterification of tetramethyl rhodamine. The analysis of the measured fluorescent intensities requires correction for fluorescence collected from outside the plane of focus of the cell and for nonpotentiometric binding of the dye. The measurements and analysis were performed on three different cell types for which there exists a body of literature on membrane potential; the potentials determined in this work were always within the range of literature values. The rhodamine esters are nontoxic, highly fluorescent dyes which do not form aggregates or display binding-dependent changes in fluorescence efficiency. Thus, their reversible accumulation is quantitatively related to the contrast between intracellular and extracellular fluorescence and allows membrane potentials in individual cells to be continuously monitored.