Optimized Ratiometric Voltage-Sensitive Dyes for Cardiac Research, Safety Pharmacology

项目摘要/摘要电位计探针 SBIR项目第二阶段的总体目标是将工具商业化， Potentiometric Probe，LLC的创始人，用于高保真光学记录电活动， 心脏细胞、组织和整个心脏。电位探针开发有机 电压敏感染料（VSD）将细胞膜上的电压变化转化为 可见的荧光变化。代替单个电极，这项技术使 使用灵敏的高速相机，其中每个像素可以被认为是电极， 大规模并行高通量筛选，或用于高分辨率时空图， 电信号传播重要的是，我们的VSD在实现双波长 电活动的比率记录;这对于心脏研究尤其重要 因为它消除了收缩引起的运动伪影，允许高保真记录的 动作电位波形人干细胞衍生的心肌细胞（hiPSC-CM）和组织 越来越多地用于研究和开发目的， 包括筛选心脏毒性的新药，将来可能用于“个性化”治疗。 医学”和患者来源细胞的分析。我们相信高通量记录 来自hiPSC-CM和组织的精确动作电位波形将是 提出的技术。最终，该技术也可以应用于临床， 心脏中动作电位传播的高分辨率成像。在这一提议中， 电位探针将合成一组稳定，无毒的比率VSD，具有广泛的 光谱窗口的选择，允许准确的记录，甚至从跳动的心脏。的 这些VSD的性能也将得到充分表征，并验证其兼容性 与敏感的细胞系进行长期和高通量记录。

Project Summary/Abstract Potentiometric Probes The overall goal of this Phase II SBIR project is to commercialize tools, invented by the founders of Potentiometric Probe, LLC, for high-fidelity optical recording of electrical activity in cardiac cells, tissues, and whole hearts. Potentiometric Probes develops organic voltage-sensitive dyes (VSDs) that convert the changes in voltage across cell membranes to visible changes in fluorescence. Instead of individual electrodes, this technology enables the use of sensitive high-speed cameras, where each pixel can be considered an electrode, for massively parallel high throughput screening, or for high-resolution spatiotemporal maps of electrical signal propagation. Importantly, our VSDs are unique in enabling dual-wavelength ratiometric recording of electrical activity; this is particularly important for cardiac studies because it eliminates contraction-induced motion artifacts, permitting high-fidelity records of the action potential waveform. Human stem cell-derived cardiomyocytes (hiPSC-CMs) and tissues derived from them are increasingly being used for research and development purposes, including screening new drugs for cardiotoxicity, and in the future may be used for “personalized medicine” and the analysis of patient-derived cells. We believe high throughput recording of accurate action potential waveforms from hiPSC-CMs and tissues will be a major market for the proposed technology. Ultimately, the technology may also be applied clinically for high-resolution imaging of action potential propagation in the heart. In this proposal, Potentiometric Probes will synthesize a set of stable, non-toxic ratiometric VSDs with a broad selection of spectral windows, allowing accurate recordings even from beating hearts. The performance of these VSDs will also be fully characterized and validated for their compatibility with sensitive cell lines for both long-term and high-throughput recordings.