Continuous cell screening using iso-dielectric separation

描述(申请人提供)：我们建议为基于细胞的筛选开发一种新的细胞分离方法。基于生物学差异的细胞分离--称为细胞筛选--是基础生物学和生物技术的基础。这些屏幕是开发治疗学和其他生物分子的重要组成部分，因此对人类健康有很大影响。对于基础生物学和生物技术，人们都对扰乱细胞--研究或改进生物合成途径--感兴趣，然后在种群中筛选理想的突变。这些理想的突变体可能会产生新的生物分子或更有效地产生现有的生物分子。为了进行筛选，人们必须有一种能够识别感兴趣的表型的实验技术。因此，使用筛子研究或设计这些有机体的总体能力从根本上取决于我们的测量能力。 特别是对于生物技术，筛选细胞以生产生物分子通常需要为每个生物分子开发一种新的分析方法，即使分离度量(例如生产)是相同的。相反，我们提出了一种新的基于生物分子生产的通用细胞分离方法，该方法是高通量、连续和实时的。这种方法利用了产生不同量生物分子的细胞之间存在的电学性质的差异。我们的假设是，大量积累的生物分子将降低细胞质的电导率和介电常数，我们可以利用这种方式来分离颗粒。我们的方法包括创建电场强度和液体电导率的比例空间梯度，这会导致电池在空间中分离，在那里它们的电学特性与液体的电学特性相匹配。我们称这种方法为等电分离(IDs)。我们对R21提案的目标是用测试颗粒和细菌来验证和表征这种方法。 这个项目与公共卫生的相关性在于，我们正在创造技术，以更好地评估产生生物分子的细胞，包括治疗学。这既有可能增加可用生物分子的种类，也有可能更有效地发现和生产它们。

DESCRIPTION (provided by applicant): We propose to develop a new approach for separating cells for cell-based screens. Separation of cells based upon biological differences-known as cell screening-is fundamental to both basic biology and biotechnology. These screens are an instrumental part of developing therapeutics and other biomolecles, and as such strongly impact human health. For both basic biology and biotechnology, one is interested in perturbing cells-to study or improve biosynthetic pathways-and then screening for desirable mutants in the population. These desirable mutants may produce new biomolecules or produce existing biomolecules more efficiently. In order to screen, one must have an experimental technique that can discern the phenotype of interest. The overall ability to study or engineer these organisms using screens is thus fundamentally dependent on our ability to measure. For biotechnology in particular, screening cells for biomolecule production typically requires development of a new assay for each biomolecule, even though the separation metric (e.g., production) is the same. Instead, we propose a new generic approach for separating cells based upon biomolecule production that is high-throughput, continuous, and real-time. This approach exploits the differences in electrical properties that exist between cells producing different amounts of biomolecule. Our hypothesis is that accumulating significant amounts of biomolecules will decrease cytoplasmic conductivity and permittivity in a way that we can exploit for particle separation. Our approach involves creating proportional spatial gradients of electric- field intensity and liquid conductivity, which causes cells to separate in space where their electrical properties match those of the liquid. We call this approach iso-dielectric separation (IDS). Our goal for this R21 proposal is to validate and characterize this approach with both test particles and bacteria. The relevance of this project to public health is that we are creating technology to better evaluate cells that produce biomolecules, including therapeutics. This has the potential to both increase the class of biomolecules available as well as more efficiently find & produce them.