SBIR Phase I: Microwell Array Plates for Dual-Mode Screening of Bead Libraries

这个小型企业创新研究(SBIR)第一阶段项目旨在开发一流的微孔阵列板，用于拥有多达100万成员的珠库的双重光学和MS成像。作为传统的基于流式细胞术的筛查的一种经济有效的替代方法，平面微珠阵列的光学成像有可能成为各种基于微珠的多路生物检测的首选分析方法。初步的实验数据表明，利用MALDI-TOF-MS可以对排列在光学透明玻璃微芯片上的微珠进行高灵敏度的测量。本项目评估的微芯片将由熔融光纤、微结构玻璃和热塑性塑料制成，以确定最佳的支撑体。微孔的独特3D结构将被用来从单个珠子中进行高度特异性的洗脱，并将它们定位在靠近微孔板表面的紧密聚焦的微点中。在不同焦距下进行的荧光成像将提供珠子结合和洗脱分析物的定量检测，而质谱仪成像将使洗脱化合物的结构表征成为可能。如果成功，该项目的更广泛的影响/商业潜力将是具有双光学和质谱学读数的微芯片的供应，这将促进高通量筛选(HTS)新方法的开发，并使质谱学能够在各种基于HTS的应用中使用。鉴于需要详细描述蛋白质组生物标记物，包括鉴定蛋白质序列变体，这种能力在生物标记物发现领域将特别重要。这项技术还将极大地惠及严重依赖HTS检测的药物发现领域。在该项目过程中将开发的高分辨率微阵列扫描技术预计将对快速增长的质谱学成像领域做出重大贡献，到目前为止，该领域主要侧重于测量各种分析物在生物组织中的分布。总体而言，拟议的工作将加快朝着单一分析平台的进展，该平台无缝地整合了荧光和质谱学，用于分析不同的微珠库，包括多肽、蛋白质和抗体微珠阵列。

This Small Business Innovation Research (SBIR) Phase I project aims to develop the first-in-class microwell array plates for dual optical and MS imaging of bead libraries comprising up to one million members. As a cost-effective alternative to the traditional flow cytometry-based screening, optical imaging of planar bead arrays has the potential to become a preferred method of analysis in a variety of multiplexed bead-based bioassays. The preliminary experimental data shows that the beads arrayed on optically transparent glass microchips also can be measured with high sensitivity by MALDI TOF MS. The microchips evaluated in this project will be fabricated from fused optic fibers, micro-structured glass and thermoplastics to determine the optimal supports. The unique 3D structure of the microwells will be utilized to perform highly specific elution of selected compounds from individual beads and their localization in tightly focused microspots near the surface of the microwell plate. Fluorescence imaging performed at varying focus distance will provide quantitative detection of the bead-bound and eluted analytes while the mass spec imaging will enable structural characterization of the eluted compounds. The broader impact/commercial potential of this project, if successful, will be the availability of microchips with dual optical and mass spec readout that will facilitate development of new methods of high-throughput screening (HTS), and enable the use of mass spectrometry in various HTS-based applications. Such ability will be particularly important in the field of biomarker discovery given the need for detailed characterization of proteomic biomarkers including identification of the protein sequence variants. This technology also will significantly benefit the field of drug discovery, which relies heavily on the HTS assays. The high-resolution microarray scanning techniques that will be developed in the course of this project are expected to make significant contribution to the rapidly growing field of mass spectrometry imaging, which so far has been focused primarily on measuring distribution of various analytes within biological tissues. Overall, the proposed work will accelerate progress toward a single analytical platform that seamlessly integrates fluorescence and mass spectrometry for the analysis of diverse libraries of microbeads including peptide, protein and antibody bead arrays.