A high-throughput assay for mitochondrial membrane potential in permeabilized yeast cells
A high-throughput assay for mitochondrial membrane potential in permeabilized yeast cells
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DOI：
10.1006/abio.2001.5139
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发表时间：
2001-06-15
影响因子：
2.9
通讯作者：
Huang, SG
中科院分区：
文献类型：
--
作者：
Farrelly, E;Amaral, MC;Huang, SG
已经开发出一种用于透化酵母细胞中线粒体膜电位的荧光测定法。该方法涉及透化质膜并测量线粒体膜电位敏感探针 3,3'-二丙基硫二碳花青碘化物 (DISC3(5); DiSC(3)) 的分布。在透化细胞中，DISC 引入通电线粒体时荧光会降低，而当线粒体膜电位被化学解偶联剂羰基氰间氯苯腙消散时，荧光会增加三到六倍。质膜电位被透化作用消除，如 K+ 和葡萄糖诱导的质膜极化缺失所示。解偶联蛋白 1 (UCP1)（一种线粒体 H+ 转运蛋白）被用作方法验证的模型。荧光强度对表达 UCP1 的细胞中的特定调节剂有强烈反应。该方法已被改编为高通量测定法，用于筛选线粒体膜电位调节剂。 (C) 2001 年学术出版社。
A fluorometric assay for mitochondrial membrane potential in permeabilized yeast cells has been developed. This method involves permeabilizing the plasma membrane and measuring the distribution of a mitochondrial membrane potential sensitive probe 3,3 ' -dipropylthiadicarbocyanine iodide (DISC3(5); DiSC(3)), In permeabilized cells, DISC, fluorescence decreased when introduced into energized mitochondria and increased three- to sixfold when the mitochondrial membrane potential was dissipated by the chemical uncoupler carbonylcyanide m-chlorophenyl hydrazone. Plasma membrane potential was abolished by permeabilization, as shown by a lack of polarization of the plasma membrane induced by K+ and glucose. Uncoupling protein 1 (UCP1), a mitochondrial H+ transporter, was used as a model for method validation. The fluorescence intensity responded vigorously to specific modulators in UCP1-expressing cells. This method has been adapted as a high-throughput assay to screen for modulators of mitochondrial membrane potential. (C) 2001 Academic Press.