Models and ultra-sensitive apparatus for electronic single cell analysis

电池是动态电子材料，其响应于环境压力而改变电子状态。已经有许多证据表明，细胞表现出与重要的生理变化相一致的介电变化，如干细胞的分化、癌细胞中多药耐药性（MDR）的发展和程序性细胞死亡（凋亡）。介电响应的单细胞测量可以识别介电亚群和这些亚群的时间变化，这是普通的散装探针不可能的。拟议的研究计划将得出定量模型的电子和生理变化之间的关系，在细胞中，也产生仪器与上级能力，以检测这些介电变化的单细胞。这项工作将集中在使用介电泳（DEP）的流动系统，使细胞从水库的分析区域，使大量的细胞在一个合理的时间范围内分析实用。与G. Bridges我开发了一种基于流动的单细胞DEP分析仪器。它可以通过检测细胞内钾离子浓度下降引起的修饰极化来检测细胞凋亡。利用微波干涉仪差示检测技术，建立了中国人卵巢细胞（CHO）凋亡的DEP细胞仪。 我建议调查一些基本的机制，介电变化的细胞中使用的化合物，是众所周知的抑制或激活特定的生化途径，将影响细胞的介电特性。例如，哇巴因抑制细胞中K+Na+泵的操作，导致细胞内离子浓度的变化，从而改变其介电性质。其他化合物控制着允许离子进出细胞的孔。这些化合物中的一些也可以证明可用作提供特定介电读出的“染料”。在上面概述的一些生理事件中，这些泵和孔被假设起主要作用。拟议的研究将测量这些化合物的影响，并推导出将特定过程的抑制与介电变化联系起来的数学模型。小体积的电池使得单电池介电测量非常具有挑战性。新的微波干涉仪设计和DEP驱动方法将用于降低介电检测极限。这项工作的目标是检测单个电池中小于5%的介电变化（克劳修斯·莫索蒂因子）。 来自这个跨学科研究计划的HQP将获得许多职业道路的技能。这个小组是少数几个有合作和工具进行这项研究的小组之一。这项工作将影响体介电和单细胞介电测量的用户和研究人员，并为单细胞生物生理学提供迫切需要的新工具。

Cells are dynamic electronic materials that change electronic state in response to environmental pressures. There have been many demonstrations that cells exhibit dielectric changes that coincide with important physiological changes such as the differentiation of stem cells, the development of multi drug resistance (MDR) in cancer cells, and programmed cell death (apoptosis). Single cell measurements of the dielectric response can identify dielectric sub-populations and temporal changes to these sub-populations that are not possible with common bulk probes. The proposed research program will derive quantitative models of the relationship between electronic and physiological changes in cells and also to produce instrumentation with superior ability to detect these dielectric changes in single cells. The work will focus on the use of dielectrophoresis (DEP) in a flow system that bring cells from a reservoir to the analysis region, making the analysis of a large number of cells practical in a reasonable time frame. In collaboration with G. Bridges I have developed a flow based single cell DEP analysis instrument. It can detect apoptosis by sensing the modified polarization that results from a drop in intracellular potassium, concentrations. Using a microwave interferometer in a differential detection approach, this DEP cytometer detected apoptosis in Chinese Hamster Ovary (CHO) cells. I propose to investigate some of the basic mechanisms of dielectric changes in cells using compounds that are well known to inhibit or activate specific biochemical pathways that will impact the dielectric properties of the cell. For example, Ouabain inhibits the operation of the K+Na+ pumps in the cell causing changes to the ion concentrations within the cell and hence its dielectric properties. Other compounds control the pores that allow ions in and out of the cells. Some of these compounds may also prove useful as “dyes” that provide a specific dielectric read out. In some of the physiological events outlined above these pumps and pores have been hypothesized to play a major role. The proposed research will measure the impact of these compounds and derive mathematical models that relate the inhibition of specific processes to dielectric changes. The small volume of cells makes single cell dielectric measurements very challenging. New microwave interferometer designs and DEP actuation methods will be used to lower the dielectric detection limits. The goal of the work is to detect dielectric changes (Clausius Mossotti Factor) of less than 5% in a single cell. The HQP from this interdisciplinary research program will gain skills for many career paths. This group is one of the few with the collaborations and tools to undertake this research. This work will impact users and researchers of bulk dielectric and single cell dielectric measurements and provide desperately needed new tools for single cell biophysiology.