Multiplexed Microarray Detection of Pluripotency Related Proteins

描述(由申请人提供)：Primorigen Biosciences将利用SBIR资金开发一种无框架、多路复用的微阵列筛选系统，用于快速识别和表征抗体匹配对，该抗体对具有对多能性和多能性至关重要的人类蛋白质的适当亲和力和特异性。该系统改进了分析内容，极大地减少了多路微阵列开发的时间和成本。这一新系统的核心是Primorigen的新型无框架微阵列设备，它允许直接在平面阵列表面针对几个结合目标对96个(或384个)样本进行多路分析。结果，可以高精度地检测数百个样本中的数十个蛋白质，每个实验产生数千个数据点，提高了筛选的速度。虽然新系统可以适用于任何蛋白质的检测，但这一第一阶段SBIR项目将专注于表征用于检测八种多能性和多能性的人类细胞标志物的分析内容，并以多路复用阵列格式对它们进行验证。第二阶段将a)完成多达15-20个多重分析的额外微阵列，b)进一步开发自动化方法以提高分析吞吐量，c)将该系统应用于与确定或维持细胞效力有关的途径分析，以及d)将分析开发扩展到内胚层、中胚层和外胚层标记。与公共健康相关：蛋白质促进了细胞的许多最基本的功能，在过去的几年里已经发现了数千种新的蛋白质序列和翻译后变体。这一蛋白质大奖加速了理解基于蛋白质的调控的努力，但研究人员需要更好的生物物理方法来定量检测蛋白质标记物。Primorigen建议用于SBIR支持的廉价、小型化、多路复用、高通量分析将满足这一需求，每个实验产生数千个蛋白质检测数据点。同时，该项目将提供新的抗体和蛋白质含量，用于检测对多能性和多能性至关重要的人类蛋白质。

DESCRIPTION (provided by applicant): Primorigen Biosciences will use SBIR funds to develop a frameless, multiplexed, microarray screening system for rapid identification and characterization of antibody matched pairs with appropriate affinity and specificity for human proteins central to pluripotency and multipotency. This system produces improved assay content and dramatically reduces the time and cost of multiplexed microarray development. The core of this new system is Primorigen's novel frameless microarray device that allows multiplexed analysis of 96 (or 384) samples against several binding targets directly on a flat array surface. As a result, dozens of proteins in hundreds of samples can be detected with high precision to generate thousands of data points per experiment, increasing speed of screening. While the new system can be adapted for detection of any protein, this Phase I SBIR project will focus on characterizing assay content for detecting eight human cell markers of pluripotency and multipotency and validating them in a multiplexed array format. Phase II will a) complete additional microarrays of up to 15-20 multiplex assays, b) further develop automated methods to increase assay throughput, c) apply this system to pathway analysis involved in determining or maintaining cell potency and d) expand assay development to endoderm, mesoderm, and ectoderm markers. PUBLIC HEALTH RELEVANCE: Proteins facilitate many of the cell's most basic functions and thousands of new protein sequences and post-translational variants have been identified in the past few years. This protein jackpot has hastened the drive to understand protein-based regulation, but researchers require better biophysical methods to quantitatively detect protein markers. Primorigen's inexpensive, miniaturized, multiplexed, high throughput assays proposed for SBIR support will address this need, generating thousands of protein detection data points per experiment. At the same time, the project will provide new antibody and protein content for assays to detect human proteins central to pluripotency and multipotency.