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

该小型企业创新研究（SBIR）第一阶段项目旨在开发一流的微孔阵列板，用于包含多达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选的具有成本效益的替代方案，平面珠阵列的光学成像具有成为多种基于多重珠的生物测定中的优选分析方法的潜力。初步的实验数据表明，排列在光学透明的玻璃微芯片上的珠也可以测量高灵敏度的MALDI TOF MS。在这个项目中评估的微芯片将由熔融光纤，微结构玻璃和热塑性塑料，以确定最佳的支持。微孔的独特3D结构将用于执行所选化合物从单个珠粒的高度特异性洗脱及其在微孔板表面附近的紧密聚焦的微点中的定位。在不同焦距下进行的荧光成像将提供珠结合和洗脱分析物的定量检测，而质谱成像将使洗脱化合物的结构表征成为可能。如果成功，该项目更广泛的影响/商业潜力将是具有双光学和质谱读数的微芯片的可用性，这将促进高通量筛选（HTS）新方法的开发，并使质谱法能够在各种基于HTS的应用中使用。鉴于需要详细表征蛋白质组生物标志物，包括鉴定蛋白质序列变体，这种能力在生物标志物发现领域将是特别重要的。这项技术也将大大有利于药物发现领域，这在很大程度上依赖于HTS分析。高分辨率的微阵列扫描技术，将在这个项目的过程中开发，预计将作出重大贡献的快速增长的质谱成像领域，到目前为止，主要集中在测量生物组织内的各种分析物的分布。总的来说，拟议的工作将加速向单一分析平台的进展，该平台无缝集成了荧光和质谱，用于分析包括肽、蛋白质和抗体珠阵列在内的各种微珠库。

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.