Novel 3-dimensional (3-D) platform for high-throughput glycomics analysis

描述(申请人提供)：多糖(碳水化合物微阵列)携带数十或数百种不同的糖以空间离散的模式固定在固体表面，在糖组学中具有广泛的潜在应用，包括快速测定碳水化合物结合蛋白的结合谱，检测用于诊断疾病的特异性抗体，表征碳水化合物细胞识别事件，以及高通量筛选抑制物以防止碳水化合物与蛋白质相互作用。然而，能够构建高质量碳水化合物微阵列以及灵敏和快速确定和表征碳水化合物-蛋白质相互作用的平台还没有跟上例如基因组和蛋白质组研究中的同等技术的步伐。为了满足这一需求，我们开发了一种独特的新型三维平台，我们已经在第一阶段研究中成功地展示了该平台用于制造高灵敏度的碳水化合物微阵列；该平台可能允许我们筛选数千种不同的碳水化合物探针与包括受体和抗体在内的蛋白质结合。在第二阶段项目中，我们建议在我们的可行性工作的基础上，开发用于高通量多糖分析的先进的碳水化合物微阵列平台。具体地说，我们建议(1)优化3D玻璃基板微阵列平台，使其能够与现有的基于荧光的多糖分析方法相结合；(2)系统比较平台表面化学对糖功能和表位展示的影响；(3)开发表位定义的高亲和力N-糖链微阵列，用于前列腺癌基于糖的生物标志物的发现；(4)开发3D SPR底物平台和无标记表面等离子体共振(SPR)碳水化合物微阵列系统。第二阶段的项目成果和交付成果将支持第二阶段的商业化目标，即实施用于碳水化合物微阵列制造的超级碳水化合物玻璃基板，以及用于碳水化合物微阵列制造的金表面等离子体共振平台基板，以实现无标记检测可用于糖链分析。后续产品将包括用于肿瘤生物标记物的定义良好的碳水化合物微阵列，以及用于高通量多糖分析的无标记SPR成像系统。平台基质和微阵列系统将为一级和二级市场带来的价值应该会吸引大公司作为合作伙伴支持商业开发、营销和分销。 与公共健康相关：碳水化合物和蛋白质之间的相互作用定义了人体内广泛的生化机制。目前，还没有商业上可行的工具来快速筛选和研究这些碳水化合物的相互作用。这项拟议的工作将开发和展示一种微阵列工具，使研究人员能够探索碳水化合物生物化学，以阐明疾病和药物机制，并识别癌症生物标记物。

DESCRIPTION (provided by applicant): Glycan (carbohydrate microarrays), carrying tens or hundreds of different sugars that are immobilized on solid surfaces in a spatially discrete pattern, possess a wide variety of potential applications in glycomics including rapid determination of the binding profile of carbohydrate-binding proteins, detection of specific antibodies for the diagnosis of diseases, characterization of carbohydrate-cell recognition events and the high-throughput screening of inhibitors to prevent carbohydrate-protein interactions. However, platforms that would enable construction of high quality carbohydrate microarrays and sensitive and rapid determination and characterization of carbohydrate-protein interactions have not kept pace with, for example, equivalent technology in genomic and proteomic research. To address this need, we have developed a unique novel 3-dimensional platform which we have demonstrated successfully for fabrication of high sensitive carbohydrate microarray in Phase I research; this platform could potentially allow us to screen thousands of different carbohydrate probes for binding to proteins including receptors and antibodies. In the Phase II project, we propose to build on our feasibility work to develop advanced carbohydrate microarray platform for high throughput glycan analysis. Specifically, we propose to (1) optimize a 3D glass-substrate microarray platform that will function with the existing fluorescent-based glycan analysis methods; (2) systematically compare of the effect of platform surface chemistry to glycan functions and epitope-displays; (3) develop epitope-defined high avidity N-glycan microarray for prostate cancer glycan-based biomarkers discovery, and (4) to develop 3D SPR substrate platform and a label-free surface plasma resonance (SPR) carbohydrate microarray system. The Phase II project results and deliverables will support the Phase II commercialization goals for implementing a Supercarbohydrate glass substrate for carbohydrate microarray fabrication that the fluorescent detection can be applied for the glycan analysis, and a Gold SPR platform substrate for carbohydrate microarray fabrication that the label-free detection can be applied for the glycan analysis. Follow-on products will include well-defined carbohydrate microarray for tumor biomarkers, and a label-free SPR imaging system for high throughput glycan analysis. The value of the platform substrates and the microarray system will bring to both the primary and secondary markets should attract large corporations as partners to support commercial development, marketing, and distribution. PUBLIC HEALTH RELEVANCE: Interactions between carbohydrates and proteins define a wide range of biochemical mechanisms in the human body. At present there is no commercially viable tool for rapid screening and investigation of these carbohydrate interactions. The proposed work will develop and demonstrate a microarray tool to allow researchers to probe carbohydrate biochemistry to elucidate disease and drug mechanisms and identify cancer biomarkers.