Ultrabright Fluorescent Nanoconstruct Enabling Widely Accessible, High Performance Multiplexed Protein Assays

摘要 生物体液和组织中多种蛋白质的检测和定量是生物学研究的基础。 生物医学研究和临床诊断的重要性，因为它是不可能理解的 复杂的，非线性的，生化系统，而不能准确地询问 件.同时询问多个蛋白质的需要在所有领域都是普遍存在的。 这是生物医学研究领域的一个主要障碍，也是充分了解健康、老龄化 疾病和对治疗干预的反应。尽管这种需求可能无处不在， 对于研究人员来说， 测量灵敏度高，动态范围大。 抗体微阵列具有简单的ELISA样工作流程，可用于测量 上千种蛋白质不幸的是，它们的灵敏度相当差， 需要使用专门且昂贵（>$75k）的读取器。这些都是重大障碍， 为大多数生物医学研究人员所采用。我们开发了一种超亮荧光 我们称之为等离子体荧光的纳米结构，比标准亮度高出5,000倍。 用于微阵列的荧光报告基因。简单地用Plasmonic Fluor代替 现有的报告子显著增加了抗体微阵列的灵敏度 工作流程的任何更改。重要的是，Plasmonic Fluor非常明亮， 微阵列可以使用更广泛可用的和通用的读取器来读取， Western blot阅读器。此外，它放宽了对阅读器仪表的要求 重要的是，这使我们能够创造一个便宜的阅读器，任何研究实验室都可以 负担得起。为了更容易地采用Plasmonic Fluor增强型微阵列，我们 还创建了微阵列数据分析软件。 在这个项目中，我们的目标是：将我们的Plasmonic Fluor的生产规模提高到早期 商业规模;更广泛地验证各种流行抗体的增强 微阵列;完成我们廉价阅读器的设计;更全面地开发我们的用户- 友好的微阵列分析软件。 我们相信Plasmonic Fluor将成为所有人的标准荧光报告分子。 微阵列，等离子体增强荧光微阵列将成为一个广泛使用的和强大的 阐明蛋白质网络在健康和疾病中的作用的工具。

Abstract Detection and quantification of multiple proteins in biological fluids and tissues is of fundamental importance in biomedical research and clinical diagnostics because it is impossible to understand complex, non-linear, biochemical systems without being able to accurately interrogate the components. The need to interrogate multiple proteins simultaneously is ubiquitous across all domains of biomedical research, and it is a major barrier to fully understanding health, ageing, disease, and response to therapeutic interventions. Though the need may be ubiquitous, there is not a widely accessible solution for researchers to make multiplexed protein measurement with high sensitivity and a large dynamic range. Antibody microarrays have a straightforward, ELISA-like workflow and can be used to measure up to thousands of proteins simultaneously. Unfortunately, they have rather poor sensitivity, and require a specialized and expensive (>$75k) reader to be used. These are significant barriers to adoption for the majority of biomedical researchers. We have developed an ultrabright fluorescent nanoconstruct we call the Plasmonic Fluor, which is >5,000X brighter than the standard fluorescent reporter used in microarrays today. Simply substituting the Plasmonic Fluor for the existing reporter significantly increases the sensitivity of antibody microarrays without requiring any change to the workflow. Importantly, the Plasmonic Fluor is so bright that it allows fluorescent microarrays to be read using more widely available and versatile readers such as a fluorescent Western blot reader. Additionally, it relaxes the requirements on the reader instrumentation significantly, which has enabled us to create an inexpensive reader that any research lab can afford. To allow even easier adoption of the Plasmonic Fluor enhanced microarrays, we have also created software for microarray data analysis. In this project, we aim to: increase the manufacturing scale of our Plasmonic Fluor to an early commercial scale; more extensively validate enhancement of a variety of popular antibody microarrays; finalize the design of our inexpensive reader; and more fully develop our user- friendly microarray analysis software. We believe the Plasmonic Fluor will become the standard fluorescent reporter molecule for all microarrays, and Plasmonic Fluor-enhanced microarrays will become a widely used and powerful tool for elucidating the role of protein networks in health and disease.