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.