Ultra-Sensitive Multiplexed Quantification of Proteins Secreted from Single-Cells - Resubmission 01

单细胞分泌蛋白质的超灵敏多重定量 这项提议旨在开发一种强大的新技术，用于多种蛋白质的超灵敏定量 由单细胞分泌。分泌的蛋白质信号协调细胞和生物体水平的功能 所有生物系统及其准确的量化对于理解生物信息是至关重要的 一般情况下，正在处理。不幸的是，信号研究由于敏感性差和局限性而受到阻碍。 当前蛋白质组学方法的多重能力，以及由于多个细胞合并而导致的结果混乱 进行总体平均分析时的读数。为了满足量化单细胞分泌的需要 跨越不同信号系统的蛋白质，我们将开发一种新的微流控技术，用于超敏感、 在单细胞水平上的多重蛋白质定量。我们将利用我们在单细胞领域的广泛专业知识 检测和微流控技术的发展和我们丰富的初步数据，以开发一个用户友好的系统 这将以一种与时间相关的方式测量来自各种细胞类型的多路蛋白质信号。 我们将通过量化包括免疫细胞在内的不同系统的信号时间进程来验证我们的方法。 癌症分泌和荷尔蒙信号。这种方法将第一次提供收集 灵敏、多重的单细胞分泌蛋白质测量，将是必不可少的 了解免疫、新陈代谢、发育和癌症中的信号。

Ultra-Sensitive Multiplexed Quantification of Proteins Secreted from Single-Cells This proposal aims to develop a powerful new technology for ultra-sensitive quantification of multiple proteins secreted from single-cells. Secreted protein signals coordinate cellular and organism-level functions across all biological systems, and their accurate quantification is crucial for understanding biological information processing in general. Unfortunately, signaling studies have been hindered by poor sensitivity and limited multiplexing ability of current proteomic methods, and the confusion of results due to pooling of multiple cell readouts when conducting population-averaged assays. To address the need to quantify single-cell secreted proteins across different signaling systems, we will develop a new microfluidic technology for ultra-sensitive, multiplexed protein quantification at the single-cell level. We will use our wide-ranging expertise in single-cell assay and microfluidic development and our extensive preliminary data to develop a user-friendly system that will measure multiplexed protein signals from a wide range of cell types, in a time-dependent manner. We will validate our method by quantifying signaling time courses of diverse systems including immune cells, cancer secretion, and hormone signaling. This method will, for the first time, provide the means to gather sensitive, multiplexed single-cell secreted protein measurements, and will be indispensable for understanding signaling in immunity, metabolism, development and cancer.