Development of SPR/MS protein array platform

描述(由申请人提供)： 这项提案的主要技术目标是开发利用表面等离子体共振(SPR)和MALDI-TOF质谱仪检测蛋白质并描绘蛋白质-蛋白质相互作用的表面等离子体共振质谱(SPR/MS)蛋白质阵列平台。在第一个可行性/中试阶段，我们将检查排列到SPR活性表面的蛋白质，阵列表面蛋白质的亲和力恢复，以及蛋白质相互作用的MALDI-TOF MS读数。通过将抗体和蛋白质定位(排列)并将其固定在芯片表面，将创建具有功能活性的蛋白质阵列。然后，蛋白质阵列将用于从溶液中亲和检索蛋白质，之后将通过MALDI-TOF质谱仪对阵列进行分析，以评估从蛋白质阵列上的斑点通过MS读出亲和捕获的蛋白质的可行性。在成功完成这些任务后，我们将进入第二个扩展开发阶段，届时将纳入高分辨率SPR检测，并将使用集成的SPR/MS蛋白质阵列平台来检测生物体液中的蛋白质和蛋白质-蛋白质相互作用。将使用高分辨率的SPR阵列仪器来显示量化阵列上的蛋白质相互作用(从单个斑点)的可行性。SPR/MS蛋白质阵列平台组件与实验控制和变量之间的接口将进一步开发和优化。如果需要，将整合更高性能的微阵列，并将评估SPR/MS蛋白质阵列平台在检测来自血浆和尿液等生物液的蛋白质和蛋白质-蛋白质相互作用方面的性能。这项开发研究的最终结果将是一个蛋白质芯片平台和方法，可用于各种蛋白质组学研究，包括高通量生物标记物分析、蛋白质-蛋白质相互作用、群体筛选努力、蛋白质治疗监测、疾病机制结构探索和诊断分析方法的发展。最终，SPR/MS蛋白质阵列平台可以使用通过微创样本收集方法获得的样本，实现对蛋白质生物标记物的快速、并行和高通量筛选，将筛选工作传播到临床和诊断实验室。

DESCRIPTION (provided by applicant): The main technical objective of this proposal is to develop Surface Plasmon Resonance Mass Spectrometry (SPR/MS) protein array platform that utilizes Surface Plasmon Resonance (SPR) and MALDI-TOF mass spectrometry for detection of proteins and delineation of protein-protein interactions. In the first feasibility/pilot phase, we will examine the protein arraying to an SPR-active surface, affinity retrieval of proteins on the array surface, and MALDI-TOF MS readout of the protein interactions. Functionally-active protein array will be created via spotting (arraying) and immobilization of antibodies and proteins onto a chip surface. The protein array will then be used for affinity-retrieval of proteins from solution, after which the array will be analyzed via MALDI-TOF mass spectrometry to gauge the feasibility of the MS readout of the affinity-captured proteins from the spots on the protein array. Upon the successful completion of these tasks, we will move into the second, expanded development phase, where high-resolution SPR detection will be incorporated and the integrated SPR/MS protein array platform will be used for detection of proteins and protein-protein interactions from biological fluids. A high resolution SPR array instrument will be employed to show the feasibility of quantification of the protein interactions (from individual spots) on the array. The interface between the components of the SPR/MS protein array platform, and the experiment controls and variables, will be further developed and optimized. If needed, a higher-performance microarrayer will be incorporated, and the performance of the SPR/MS protein array platform in detection of proteins and protein-protein interactions from biological fluids such as plasma and urine will be evaluated. The final result of this developmental research will be a protein chip platform and methods that can be employed into various lines of proteomics research, including high-throughput biomarker analysis, protein-protein interactions, population screening efforts, therapeutic monitoring of proteins, exploration of disease mechanism structures, and diagnostic assays development. Ultimately, the SPR/MS protein array platform could enable rapid, parallel, and high-throughput screening of protein biomarkers, using samples obtained through minimally invasive sample collection methods, propagating the screening efforts into the clinical and diagnostic laboratories.