SBIR Phase II: Universal Proteome Sample Preparation Kit Development

这个小型企业创新研究（SBIR）第二阶段项目的更广泛的影响/商业潜力是开发一种通用的蛋白质样品制备试剂盒，从任何生物样品中提取蛋白质，去除污染物，并为下游分析方法做好准备，其产量和覆盖范围比目前可用的技术更高。 蛋白质是细胞的分子机器，它完成了保持人类健康所需的所有工作，蛋白质分析的结合对于发展对集成生物系统的清晰理解，实现精准医学和快速药物发现的承诺变得越来越重要。样品制备是任何基于蛋白质的项目工作流程的第一步，如果没有高产、快速和可重复的样品制备方法，下游分析难以解释且高度可变，导致项目时间长和资源浪费。 拟议的创新提高了样本产量、重现性和速度，加速了寻求发现新疗法和诊断方法的研究，并提高了成本效益。 它还将从同一样品中纯化其他重要的生物分子，如DNA，RNA和代谢物，使研究人员能够从每个样品中获得更多信息，在制备一个样品所需的时间内制备多个样品，并通过使用相同的起始材料提高重现性。拟议的项目将利用可逆的蛋白质标记化学来创建一个单管工作流程，用于捕获、洗涤和洗脱蛋白质样品进行分析。 由于这种新型可逆的蛋白质标记化学不依赖于催化剂，不产生副产物，并且完全是双正交的，因此它具有与所有类型的缓冲系统通用的潜力，包括通常用于细胞裂解和蛋白质溶解的苛刻变性剂和去污剂。 这项拟议的工作将优化利用这种样品制备技术在最常用的缓冲系统，模式生物，并将显示上级产量在诊断样品类型，如血液，尿液和脑脊液的制备。 此外，由于我们的可逆蛋白质标签不会与其他生物分子发生交叉反应，我们将证明我们也可以使用这种技术来纯化其他重要的生物分子，如DNA，RNA和代谢物。 该项目开发的技术将彻底改变研究人员制备分析样品的方式，为他们提供模块化工具，可以从相同的起始样品中纯化他们需要的任何生物分子，减少时间，提高产量，该奖项反映了NSF的法定使命，并通过使用基金会的知识价值和更广泛的影响审查标准。

The broader impact/commercial potential of this Small Business Innovation Research (SBIR) Phase II project is to develop a universal protein sample preparation kit that extracts proteins from any biological sample, removes contaminants, and prepares them for downstream analytical methods with higher yields and deeper coverage than currently available technology. Proteins are the molecular machines of the cell that do all the work required to keep humans healthy, and incorporation of protein analysis is becoming increasingly important for developing a clear understanding of integrated biological systems, realizing the promise of precision medicine, and rapid drug discovery. Sample preparation is the first step in any protein-based project workflow and without a high-yield, fast, and reproducible way to prepare samples, downstream analyses are difficult to interpret and highly variable, leading to long project times and wasted resources. The proposed innovation improves sample yields, reproducibility, and speed, accelerating research seeking to discover new therapies and diagnostics and improving cost-effectiveness. It will also purify other important biomolecules from the same sample, such as DNA, RNA, and metabolites, allowing researchers to obtain more information from each sample, prepare multiple samples in the time it takes to prepare one, and improve reproducibility by using the same starting material. The proposed project will utilize reversible protein tagging chemistry to create a one-tube workflow for the capture, wash, and elution of protein samples for analysis. Because this novel reversible protein tagging chemistry is not dependent on a catalyst, does not produce byproducts, and is completely biorthogonal, it has the potential to work universally with all types of buffer systems, including harsh denaturants and detergents commonly used for cell lysis and protein solubilization. This proposed work will optimize the utilization of this sample preparation technology in the most used buffer systems, model organisms, and will show superior yields in the preparation of diagnostic sample types such as blood, urine, and cerebrospinal fluid. Additionally, because our reversible protein tag does not cross react with other biomolecules, we will show that we can also use this technology to purify other important biomolecules such as DNA, RNA, and metabolites. The technology developed in this project will revolutionize how researchers prepare samples for analysis, providing them with modular tools that can purify any biomolecules they need from the same starting sample, reducing time, improving yields, and decreasing batch to batch variability by comparing within the same starting sample.This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.