Collaborative Research: Plasmonic Nanoantenna Electrode Arrays (NEAs) for Massively Multiplexed Identification of Stem-Cell Derived Cardiac Cells in Regenerative Therapies

合作研究：等离子体纳米天线电极阵列（NEAs）用于再生治疗中干细胞来源的心脏细胞的大规模多路识别非技术摘要：心脏病是美国主要的死亡原因之一。基于干细胞的再生疗法是最有前途的治疗技术之一。然而，干细胞衍生的细胞并不统一；最初的细胞培养中只有一定比例的细胞发育成我们感兴趣的细胞类型。留在细胞群中的未分化细胞可能导致肿瘤。此外，未成熟的细胞或过度敏感的细胞会阻碍心肌细胞的同步跳动，从而导致心力衰竭。目前检测基于干细胞的心脏细胞纯度的方法依赖于细胞表面标记物，这不是确定细胞功能的精确方法。该建议提供了一种高通量筛选技术，通过心肌细胞在收缩过程中特定的膜电位变化直接测量分化心肌细胞的功能。提出的分子纳米等离子体无标签电压传感器将允许筛选融合细胞培养中的单个细胞膜电位，并提供从混合组中选择和纯化功能细胞的准确方法。这种精确技术的发展将是基于干细胞的心脏再生研究和策略的一个显著的技术飞跃。除了科学和技术的进步，这项研究计划将提供教育机会，代表性不足的群体和少数民族，并加强参与纳米科学和技术的本科生和研究生。技术摘要：本研究的目的是引入超灵敏的分子等离子体电压探针，用于无创、实时和亚细胞的心脏细胞膜电位精确测绘。这些电生理纳米探针可以通过大规模平行和精确地绘制单细胞膜电位，对干细胞来源的心脏细胞的分化产生重大影响。鉴于缺乏具有高空间和时间精度的实验电生理技术，该研究计划可以为心脏细胞研究和再生治疗做出重大贡献。本研究计划的具体目标是：(1)通过电磁模拟、高通量制造和化学合成技术以及光学/电学表征来开发分子等离子体电压传感器。(2)以微秒级时间分辨率和高信噪比实现对衍射极限光斑尺寸下微小电位变化的实时、无标签检测。(3)实现大细胞群中单细胞的无损成像，区分培养/多细胞状态下的单个细胞特征。提出的研究计划涉及分子等离子体器件的理论认识和数值设计。融合纳米/微米元件的器件将采用光刻技术和合成技术制造。制造的设备将使用具有不同密度和细胞成分的可兴奋细胞群进行测试。此外，从hiPSC分化的心肌细胞的膜电位变化将被实时测量。

Collaborative Research: Plasmonic Nanoantenna Electrode Arrays (NEAs) for Massively Multiplexed Identification of Stem-Cell Derived Cardiac Cells in Regenerative TherapiesNontechnical Abstract: Heart diseases are one of the leading causes of death in the US. Stem cellbased regenerative therapies are among the most promising treatment techniques. However, cells derived from stem cells are not uniform; only some percentage of the initial cell culture develops into the cell type of interest. Undifferentiated cells that remain within the cell population could lead to tumor.Furthermore, immature cells or cells with over-sensitivity would hinder the synchronous beating of the heart muscle cells, which can cause heart failure. Current methods to examine the purity of stem-cell based heart cells depend on cell surface markers, which is not a precise way to determine cellular functionality. This proposal offers a high-throughput screening technique to directly measure thefunctionality of differentiated heart muscle cells through their specific membrane potential changes during contraction. The proposed molecular-nanoplasmonic label-free voltage sensors will allow screening of single cell membrane potentials within confluent cell cultures and provide an accurate method for selecting and purifying functional cells from a mixed group. Development of such a precise technique would present a remarkable technological leap in stem cell-based research and strategies for cardiac regeneration. In addition to scientific and technological advancements, this research program will provide educational opportunities to underrepresented groups and minorities, and enhance involvement of undergraduate and graduate students in nanoscience and technology.Technical Abstract: The objective of this research proposal is to introduce ultrasensitive molecularplasmonic voltage probes for non-invasive, real-time and subcellular precision mapping of cardiac cell membrane potentials. These electrophysiological nanoprobes could have significant impact in differentiation of stem cell derived cardiac cells through massively parallel and precise mapping of singlecell membrane potentials. Given the lack of experimental electrophysiological techniques with high spatial and temporal precision capabilities, this research program could significantly contribute to cardiac cell studies and regenerative therapies. The specific objectives of this research program are:(1) to develop molecular-plasmonic voltage sensors by using electromagnetic simulations, high throughput fabrication and chemical synthesis techniques, and optical/electrical characterization.(2) to realize real time and label free detection of tiny potential variations at diffraction limited spot sizes with microsecond temporal resolutions and high signal-to-noise ratios.(3) to achieve non-destructive imaging of single cells in large cell populations and distinguish individual cell characteristics in -cultured/multiple cell state.The proposed research program involves theoretical understanding and numerical design of molecularplasmonic devices. Devices merging nano/micro-meter components will be fabricated using state of lithography and synthesis techniques. Fabricated devices will be tested using excitable cell populations with varying densities and cell compositions. Furthermore, changes in the membrane potentials ofcardiomyocytes that are being differentiated from hiPSC will be measured in real-time.