Voltage‐sensitive cyanine dye fluorescence signals in lymphocytes: Plasma membrane and mitochondrial components
Voltage‐sensitive cyanine dye fluorescence signals in lymphocytes: Plasma membrane and mitochondrial components
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淋巴细胞中的电压敏感花青染料荧光信号：质膜和线粒体成分
DOI：
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发表时间：
1985
影响因子：
5.6
通讯作者：
T. Chused
中科院分区：
文献类型：
--
作者：
H. A. Wilson;B. Seligmann;T. Chused
使用氧杂和吲哚羰花菁二-O-C5（3）研究了小鼠和人外周血白细胞中花菁染料荧光信号的来源。和di-l-C5（3）。在经修改以允许同时进行四参数分析（包括双色荧光或荧光偏振测量）的流式细胞仪中测量来自用纳摩尔浓度的染料悬浮的单个细胞的荧光信号。发现线粒体膜电位（Km）和质膜电位（Km）对总电压敏感荧光信号的贡献取决于平衡的细胞外染料浓度，在这些实验中通过改变染料与细胞密度的比率来操纵。因此，可以选择放大λ m或λ pm组分的条件。混合细胞悬液中淋巴细胞或多形核白细胞（PMN）的选择性去极化表明，确定细胞和培养基之间的染料分区是评估细胞反应的异质性所必需的花青染料荧光。在细胞外染料浓度超过5纳米平衡的细胞悬浮液中，线粒体和质膜染料毒性观察。在小鼠脾淋巴细胞中，质膜毒性（染料诱导的去极化）对B淋巴细胞具有选择性。某些问题，在低染料浓度和线粒体抑制剂的干扰下，缬氨霉素校准的pH4.5。这些研究结果解决了目前的争议有关在完整的细胞中的花青染料荧光法测量的pH2.5和pH2.5。在低花菁染料浓度下的选择性毒性的发现表明，细胞间静息细胞的差异或细胞活化时细胞间静息细胞的变化可能反映了对染料毒性的可变易感性，而不是内在的细胞特性。
The origin of the cyanine dye fluorescence signal in murine and human peripheral blood leukocytes was investigated using the oxa‐ and indo‐carbo‐cyanines di‐O‐C5(3). and di‐l‐C5(3). Fluorescence signals from individual cells suspended with nanomolar concentrations of the dyes were measured in a flow cytometer modified to permit simultaneous four‐parameter analysis (including two‐color fluorescence or fluorescence polarization measurements). The contributions of mitochondrial membrane potential (Ψm) and plasma membrane potential (Ψpm) to the total voltage‐sensitive fluorescence signal were found to depend on the equilibrium extracellular dye concentration, manipulated in these experiments by varying the ratio of dye to cell density. Hence, conditions could be chosen that amplified either the Ψm or the Ψpm component. Selective depolarization of lymphocytes or polymorphonuclear leukocytes (PMN) in mixed cell suspensions demonstrated that defining the partition of dye between cells and medium is requisite to assessing the heterogeneity of cell responses by cyanine dye fluorescence. At extracellular dye concentrations exceeding 5 nM in equilibrated cell suspensions, both mitochondrial and plasma membrane dye toxicity were observed. In murine splenic lymphocytes, plasma membrane toxicity (dye‐induced depolarization) was selective for the B lymphocytes. Certain problems in calibration of Ψpm with valinomycin at low dye concentrations and perturbations of Ψpm by mitochondrial inhibitors are presented. These findings address the current controversy concerning Ψm and Ψpm measurement in intact cells by cyanine dye fluorescence. The finding of selective toxicity at low cyanine dye concentrations suggest that purported differences in resting Ψm among cells or changes in Ψpm with cell activation may reflect varible susceptibility to dye toxicity rather than intrinsic cell properties.
影响因子：
64.8
作者：
DECOURSEY, TE;CHANDY, KG;CAHALAN, MD
通讯作者：
CAHALAN, MD
T 淋巴细胞胆碱能受体的电生理学。
DOI：
10.1073/pnas.77.7.4317
发表时间：
1980
影响因子：
11.1
作者：
Shapiro,HM;Strom,TB
通讯作者：
Strom,TB