Plasmonic Assay for Detection of Biomarkers at the Single-Molecule Level

描述（由申请人提供）：本提案的目标是开发一种敏感，高通量的局部表面等离子体共振检测方法，用于检测范围广泛的生物标志物，其水平低至单分子水平，并具有高度可扩展的动态范围。我们最近的研究表明，等离子共振纳米粒子对它们与附近导电衬底的接近度非常敏感。它们在共振（或颜色）中显示出较大的光谱位移，以及在与附近金属薄膜的距离发生纳米级变化时散射光偏振的变化。此外，由于易于功能化和检测，这种纳米颗粒也是生物分子的有效标记。这里提出了一种方案，即等离子共振纳米粒子和金属表面功能化与捕获分子设计，以识别特定的生物标志物大小不等，从小分子/DNA目标到蛋白质。暴露于临床样品后，功能化纳米颗粒结合目标分析物，然后定向到由互补捕获分子的表面模式定义的导电基底上的特定区域。本文提出的全新创新是，通过在数字图像中搜索预定的纳米颗粒颜色和/或散射极化，结合到纳米颗粒上的目标将被检测出来，并与没有目标的纳米颗粒的非特异性吸附区分开来，这与结合目标决定的物理尺寸（因此，纳米颗粒和薄膜之间的间距）是特定的。本建议的具体目的如下：传感器响应的优化。2. “受体”功能化NPs的设计和传感器芯片的制造。3. 检测临床分析。公共卫生相关性：随着我们对疾病知识的扩展，可靠地检测各种大小和浓度的生物分子的需求变得更加重要。这项工作的目标是开发一种新的检测平台，可以从单分子水平扩展到多路分析水平，其中信号很容易从背景噪声中伪装出来，从而提高临床生物标志物分析的准确性、灵敏度和适用性。

DESCRIPTION (provided by applicant): The goal of this proposal is to develop a sensitive, high-throughput localized surface plasmon resonance assay for the detection of a wide range of biomarkers at levels reaching down to that of single molecules and with a highly scalable dynamic range. We have recently shown that plasmon resonant nanoparticles are highly sensitive to their proximity to nearby conductive substrates. They display large spectral shifts in resonance (or color) as well as changes in scattered light polarization upon nanometer-scale changes in distance from a nearby metal film. In addition, such nanoparticles are also effective labels for biomolecules because of their ease of functionalization and detection. Proposed here is a scheme whereby plasmon resonant nanoparticles and metal surfaces are functionalized with capture molecules designed to recognize specific biomarkers varying in size from small molecules/DNA targets to proteins. Upon exposure to clinical samples, the functionalized nanoparticles bind target analytes and are then directed to specific areas on conductive substrates defined by surface patterns of complimentary capture molecules. The completely novel innovation proposed here is that targets bound to nanoparticles will be detected and discriminated from nonspecific adsorption of nanoparticles without targets by searching in a digital image for a predetermined nanoparticle color and/or scattering polarization that is specific to the physical size (and hence, spacing between nanoparticle and film) determined by bound target. Specific aims of this proposal are as follows: 1. Optimization of the sensor response. 2. Design of "receptor" functionalized NPs and fabrication of sensor chips. 3. Detect clinical analytes. Public Health Relevance: As our knowledge of disease expands the need to reliably detect biomolecules of various sizes and concentrations becomes more important. The goal of this work is to develop a novel detection platform that is scalable from the single molecule level to the multiplexed assay level where signal is easily disguishable from background noise, thus increasing the accuracy, sensitivy, and applicability of clinical biomarker assays.