Development of SPR/MS protein array platform

本提案的主要技术目标是开发表面等离子体共振质谱 (SPR/MS）蛋白质阵列平台，其利用表面等离子体共振（SPR）和MALDI-TOF质谱 用于检测蛋白质和描绘蛋白质-蛋白质相互作用的光谱法。在第一个可行性/试点中 阶段，我们将研究蛋白质阵列到SPR活性表面，亲和修复蛋白质上的蛋白质， 阵列表面，以及蛋白质相互作用的MALDI-TOF MS读数。功能活性蛋白质阵列将 通过将抗体和蛋白质点样（排列）和固定到芯片表面上来产生。的 然后，蛋白质阵列将用于从溶液中亲和回收蛋白质，之后，将该阵列 通过MALDI-TOF质谱分析，以衡量MS读出的可行性。 从蛋白质阵列上的点亲和捕获的蛋白质。在圆满完成这些任务后， 我们将进入第二个扩展的开发阶段，在这个阶段，高分辨率的SPR检测将是 整合的SPR/MS蛋白质阵列平台将用于检测蛋白质， 蛋白质与蛋白质之间的相互作用。将采用高分辨率SPR阵列仪器， 显示了在阵列上定量蛋白质相互作用（来自单个点）的可行性。的 SPR/MS蛋白质阵列平台组件与实验对照之间的接口， 变量，将进一步开发和优化。如果需要的话，一个更高性能的微阵列将是 结合，和SPR/MS蛋白质阵列平台在蛋白质检测中的性能， 将评价来自生物流体如血浆和尿的蛋白质-蛋白质相互作用。最终结果 这项发展研究的一部分将是一个蛋白质芯片平台和方法，可以用于各种 蛋白质组学研究，包括高通量生物标志物分析，蛋白质-蛋白质相互作用， 群体筛选工作，蛋白质的治疗监测，疾病机制结构的探索， 和诊断分析开发。最终，SPR/MS蛋白质阵列平台可以实现快速， 平行和高通量筛选蛋白质生物标志物，使用通过最低限度地 侵入性样本采集方法，将筛查工作推广到临床和诊断领域， laboratories.

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.