Validation of acoustic tweezers for single-cell analyses of purine metabolism

 产品说明：缺乏一种可以同时实现高通量、高精度和高细胞完整性的单细胞操作技术是细胞间通讯研究的主要障碍。最近，我们的跨学科团队开发了一种基于表面声波（SAW）的微流体平台，称为“声学镊子”，与现有的单细胞分析细胞操作技术相比具有显着优势。我们的声学镊子平台能够以亚微米精度调节单个细胞之间的距离。此外，它具有高度可扩展性，能够为高通量研究创建大量的细胞排列。细胞不需要标记，可以在其天然培养基中培养。此外，用于操纵细胞的声功率和频率与超声成像中使用的范围相同，这已被证明具有高度生物相容性。最后，SAW产生所需的组件小而便宜，并且设备本身易于操作。凭借这些优势，声镊在其以高通量方式在单细胞水平上提供细胞内通信的精确时空控制同时保持细胞完整性的能力方面具有开创性。声学镊子的变革潜力已经在几种同型和异型细胞群体中的间隙连接介导的功能性细胞间通讯的研究中得到了证明，通过可视化细胞之间的荧光染料转移。我们在这个项目中的目标是进行声学镊子的先进开发，并验证它们在细胞内代谢途径的细胞间通讯的影响的研究。因此，我们将致力于以下几个方面的研究：（1）在单细胞水平上高产量、高通量地表征细胞间通讯和嘌呤体组装的声镊的发展;（2）利用声镊在原代细胞模型中对嘌呤体组装的多参数研究;和（3）使用声镊在神经元模型中进行嘌呤体组装和嘌呤代谢的单细胞分析。在拟议的项目完成后，我们希望以Lesch-Nyhan病（LND）为疾病模型，以嘌呤体为代谢状态的指标，揭示基因型如何影响复杂表型的机制。由于其独特的能力，以高通量创建具有规定架构的多细胞组件，我们预计声学镊子将成为单细胞分析的宝贵工具，并将满足生物工程，生物医学和制药研究领域的许多未满足的需求。

 DESCRIPTION: The lack of a single-cell manipulation technique that can simultaneously achieve high throughput, high precision, and high cell integrity is a major roadblock for studies of intercellular communication. Recently, our interdisciplinary team has developed a surface acoustic wave (SAW)-based microfluidic platform called "acoustic tweezers" that possesses significant advantages over existing cell-manipulation techniques for single-cell analysis. Our acoustic tweezers platform is able to modulate the distances between individual cells with sub-micron precision. In addition, it is highly scalable and capable of creating a large array of celluar arrangements for high-throughput studies. Cells do not need to be labelled and can be cultured in their native media. Furthermore, the acoustic power and frequency used to manipulate cells are in the same range as those used in ultrasonic imaging, which has proven to be highly biocompatible. Finally, the components required for SAW generation are small and inexpensive, and the device itself is easy to operate. With these advantages, the acoustic tweezers are groundbreaking in their ability to provide precise spatiotemporal control of intracellular communication at the single-cell level in a high-throughput manner while preserving cell integrity. The transformative potential of acoustic tweezers has already been demonstrated in studies on gap junction-mediated functional intercellular communication in several homotypic and heterotypic cell populations by visualizing the transfer of fluorescent dyes between cells. Our objective in this project is to conduct advanced development of acoustic tweezers and validate them in studies on the effects of intercellular communication on metabolic pathways within the cell. We will, therefore, pursue the following specific aims: (1) advanced development of acoustic tweezers for high-yield, high-throughput characterization of intercellular communication and purinosome assembly at the single-cell level; (2) multi-parametric investigation of purinosome assembly in a primary cell model using acoustic tweezers; and (3) single-cell analyses of purinosome assembly and purine metabolism in a neuronal model using acoustic tweezers. At the completion of the proposed project, we hope to uncover the mechanism for how a genotype affects complex phenotype using Lesch-Nyhan disease (LND) as the disease model and purinosome as an indicator of metabolic state. Due to its unique ability to create multicellular assemblies with prescribed architectures in high throughput, we expect that the acoustic tweezers will become an invaluable tool for single-cell analysis and will fulfill many unmet needs in the bioengineering, biomedical, and pharmaceutical research communities.