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

描述(申请人提供)：这项建议的目标是开发一种灵敏的、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记物，其水平可以精确到单分子水平，并且具有高度可扩展的动态范围。我们最近已经证明，等离子激元共振纳米粒子对其与附近导电衬底的接近高度敏感。它们表现出共振(或颜色)的巨大光谱位移，以及与附近金属膜之间的纳米级距离变化时散射光偏振的变化。此外，这种纳米粒子也是生物分子的有效标记物，因为它们易于功能化和检测。这里提出了一种方案，利用捕获分子对等离子激元共振纳米颗粒和金属表面进行功能化处理，以识别从小分子/DNA靶标到蛋白质的不同大小的特定生物标志物。当接触到临床样本时，功能化的纳米颗粒与目标分析物结合，然后被定向到导电基质上由互补捕获分子的表面图案定义的特定区域。这里提出的全新的创新是，通过在数字图像中搜索预定的纳米颗粒颜色和/或特定于由结合目标确定的物理尺寸(因此，纳米颗粒与膜之间的间距)的散射偏振，结合到纳米颗粒上的目标将被检测到，并与没有目标的纳米颗粒的非特异性吸附区分开来。该方案的具体目标如下：1.优化传感器响应。2.“受体”功能化纳米粒子的设计和传感器芯片的制作。3.检测临床分析物。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型的检测平台，该平台可以从单分子水平扩展到多重检测水平，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记物分析的准确性、敏感度和适用性。

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.