Maher Michael P, Wu Nyan-Tsz, Ao Hong
Johnson & Johnson Pharmaceutical Research and Development L.L.C., San Diego, California 92121, USA.
J Biomol Screen. 2007 Aug;12(5):656-67. doi: 10.1177/1087057107302113. Epub 2007 May 21.
Many high-throughput ion channel assays require the use of voltage-sensitive dyes to detect channel activity in the presence of test compounds. Dye systems employing Förster resonance energy transfer (FRET) between 2 membrane-bound dyes are advantageous in combining high sensitivity, relatively fast response, and ratiometric output. The most widely used FRET voltage dye system employs a coumarin fluorescence donor whose excitation spectrum is pH dependent. The authors have validated a new class of voltage-sensitive FRET donors based on a pyrene moiety. These dyes are significantly brighter than CC2-DMPE and are not pH sensitive in the physiological range. With the new dye system, the authors demonstrate a new high-throughput assay for the acid-sensing ion channel (ASIC) family. They also introduce a novel method for absolute calibration of voltage-sensitive dyes, simultaneously determining the resting membrane potential of a cell.
许多高通量离子通道检测需要使用电压敏感染料来检测受试化合物存在时的通道活性。采用两种膜结合染料之间的荧光共振能量转移（FRET）的染料系统具有高灵敏度、相对快速响应和比率输出的优势。使用最广泛的FRET电压染料系统采用一种香豆素荧光供体，其激发光谱依赖于pH值。作者已经验证了一类基于芘部分的新型电压敏感FRET供体。这些染料比CC2-DMPE亮得多，并且在生理范围内对pH不敏感。利用这种新的染料系统，作者展示了一种针对酸敏感离子通道（ASIC）家族的新型高通量检测方法。他们还引入了一种对电压敏感染料进行绝对校准的新方法，同时测定细胞的静息膜电位。
