Metal Enhancement for Near-IR Fluorescence Based Membrane Assays

描述(申请人提供)：位于银纳米颗粒附近的荧光分子产生更强的荧光强度，同时增加光稳定性和缩短寿命。人们对这些银结构进行了非常详细的研究，因为它们在沉积到固体表面(如玻璃或石英)时能够增加荧光强度。然而，关于银纳米结构增强沉积在多孔膜表面的荧光团的能力的信息很少。这项提议的总体目标是利用金属增强荧光来增加膜基质上的近红外荧光发射。基于膜的分析，如免疫印迹，通常用于临床环境，以确认其他筛查测试所建议的严重诊断，并用于研究环境，以表征和定量感兴趣的蛋白质。与可见光光谱中产生的背景相比，近红外中基于荧光的分析提供了显著降低背景的额外优势，有助于降低检测极限。从长远来看，这项工作应该适用于以将荧光染料捕获在膜衬底上的方式利用荧光的大多数系统。膜表面的近红外金属增强荧光应该提供一种可靠和有效的方法来增加荧光发射和降低检测极限，提供一种与当前蛋白质组和基因组应用相兼容的工具。未来的努力可能会使用金属增强荧光来提高基于膜的DNA、蛋白质和膜上抗体阵列的可检测性。该项目致力于开发一种用于蛋白质和斑点印迹分析的增强剂，它将低本底近红外锂染料与金属增强荧光(MEF)的使用相结合，以产生更灵敏的基于膜的分析方法，能够将检测限降低一个数量级。这些蛋白质印迹和斑点印迹分析在灵敏度方面的进展将有助于分析分子、细胞和细胞间的过程，以及这些过程之间的相互关系。

DESCRIPTION (provided by applicant): Fluorescent molecules that are positioned in close proximity to silver nanoparticles produce increased fluorescence intensity along with increased photostability and decreased lifetimes. These silver structures have been studied in great detail for their ability to increase fluorescent intensities when deposited onto solid surfaces, such as glass or quartz. Little information exists, however, on the ability of silver nanostructures to enhance fluorophores deposited onto porous membrane surfaces. The overall objective of this proposal is to use metal enhanced fluorescence to increase near- infrared fluorescence emissions on membrane substrates. Membrane based assays, such as western blots are routinely used in clinical settings to confirm serious diagnoses suggested by other screening tests, and in research settings to characterize and quantitate proteins of interest. Fluorescence based assays in the near-infrared offer the additional advantage of significantly lowering the background compared to that generated in the visible spectrum, helping to lower the limit of detection. In the long term, this work should be applicable to most systems that utilize fluorescence in a manner that traps the fluorescent dyes on a membrane substrate. Near-infrared metal enhanced fluorescence on membrane surfaces should provide a reliable and efficient procedure to increase fluorescent emissions and decrease the limit of detection, providing a tool that is compatible with current proteomic and genomic applications. Future endeavors could use metal enhanced fluorescence to increase the detectability of membrane based DNA, protein, and antibody arrays on membranes. This project focuses on developing an enhancement reagent for Western and dot blot analyses, which integrates the use of low background near-infrared LI-COR dyes with Metal Enhanced Fluorescence (MEF), to produce more sensitive membrane based assays capable of reducing the limit of detection by an order of magnitude. Advances in sensitivity for these Western and dot blot assays will aid in the analysis of molecular, cellular, and inter-cellular processes, and the relationship of these processes with respect to one another.