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

项目摘要/抽象电位测头 这个第二阶段SBIR项目的总体目标是将由 用于高保真光学记录电活动的电位计探头LLC的创始人 心肌细胞、组织和整个心脏。电位法探头显影有机 电压敏感染料(VSD)，将细胞膜上的电压变化转换为 可见的荧光变化。这项技术不是单独的电极，而是使 使用灵敏的高速摄像机，其中每个像素都可以被视为电极，用于 大规模并行高通量筛选，或用于高分辨率时空地图 电信号传播。重要的是，我们的VSD在支持双波长方面独一无二 电活动的比率记录；这对心脏研究特别重要 因为它消除了收缩引起的运动伪像，允许高保真地记录 动作电位波形。人干细胞来源的心肌细胞和组织 从它们衍生出来的越来越多地被用于研究和开发目的， 包括筛选心脏毒性的新药，并在未来可能被用于“个性化” 医学“和病人来源细胞的分析。我们相信高通量记录 来自HiPSC-CMS和组织的准确动作电位波形将是 建议的技术。最终，这项技术也可能应用于临床 心脏动作电位传播的高分辨率成像。在这份提案中， 电位法探针将合成一套稳定、无毒的比例式VSD，具有广泛的 选择光谱窗口，即使是跳动的心脏也能进行准确的记录。这个 还将对这些VSD的性能进行全面表征，并验证其兼容性 具有敏感的细胞系，可用于长期和高通量记录。

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.