CAREER: microMOSAIC Frameworks for Next-Generation Proteomic Technology

职业：下一代蛋白质组技术的 microMOSAIC 框架

• 批准号: 1056035
• 负责人: Amy Herr
• 金额: $ 33.25万
• 依托单位: University of California-Berkeley
• 依托单位国家: 美国
• 项目类别: Continuing Grant
• 财政年份: 2011
• 资助国家: 美国
• 起止时间: 2011-02-15 至 2017-01-31
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=1056035&HistoricalAwards=false
• 关键词: CAREER; microMOSAIC; Frameworks; Next; Generation

This research will introduce novel optically patterned microchamber and flow control technology to overcome integration barriers that have prevented two decades of microfluidics research from offering compelling alternatives to the work-horse applications? Western blotting and two-dimensional electrophoresis (2DE). Breakthroughs in proteomic technology are essential: while the genomic revolution has had sweeping impact on understanding of life processes, the arguably more important ?proteomic revolution remains unrealized. Proteins are more directly linked with function, more biochemically complex, and 10-100x more numerous than genes. Thus, nearly all life science studies require multi-step assays such as Western blot and 2DE. Yet, benchtop multi-step separations demand huge labor, time investment, and precious sample to yield semi-quantitation; these assays would benefit immensely from integration. The unique ìMosaic integration strategy--microchambers regionally photopatterned with discrete nanostructured separation materials--has the potential to revolutionize multi-step separations for the pressing proteomic challenges of the 21st century.Multi-step proteomic assays measure not one, but multiple physicochemical and functional properties. To date, the performance of multi-step microseparations has suffered from status quo design strategies that couple a first assay step (a single microchannel) to a second assay step that is an array of microchannels. Regrettably this approach ?discretizes? first-stage separation readouts by mapping them to discrete compartments in a second-stage. To overcome integration losses, the PI proposes a radical departure from status quo by introducing microchambers regionally photopatterned with discrete nanomaterials. Fundamental studies underpinning ìMosaic integration technology will include: 1) Development & quantitative evaluation of micro/nanofabrication techniques and devices for charge-, chemical-, and photo-patterning of polymers to achieve localized biochemical/physical function within fluidic networks. Reactions within the patterned nanostructured materials will be studied using fluorescence imaging, determining specificity, kinetics, and overall affinity using controlled, homogenous reagent flows in control experiments. 2) Separation integration with dispersion control strategies for high performance multi-step separations. PI will investigate the interplay between electromigration, diffusion, and reaction empirically, analytically, and with 3D multiphysics solvers. The PI will identify and control transport and reaction mechanisms; i.e., band broadening with spatially non-uniform surface reactions. Although coupled dispersion and reaction are understood in homogeneous microchannel networks, such is not the case with 3D geometries, 3D heterogeneous reaction patterns underpinning the ìMosaic multi-step assays proposed here. Together, these studies will provide new knowledge in: fabrication; nanostructured materials, separations device and assay engineering; experimental methods. While available bench-top proteomic tools are ubiquitous, significant limitations in throughput, quantitation, dynamic range, and automation exist. The proposed novel and widely applicable tools will specifically advance each ? impacting systems biology, synthetic biology, and biofuels. Likely outcomes include: simulation-based screening of therapeutic agents, high-throughput identification of new cellulosic biofuels, and rational selection algorithms for bacterial bioremediation. The PI will integrate research with teaching through new multi-media lecture material in two developing courses and as hands-on modules in a new K-12 partnership (Lawrence Hall of Science Ingenuity Labs). The PI proposes a comprehensive strategy to recruit and retain underrepresented students and holds leadership positions on three NSF REU?s. As faculty advisor to Berkeley?s Society of Women Engineers (SWE), she will bolster SWE and Berkeley?s nascent Graduate Women in Engineering group through NSF support. She is a leader in the larger technical community through conference chair-ship, technical program committee service, and community building (e.g., Women in MEMS).

这项研究将引入新的光学图案化微室和流量控制技术，以克服集成障碍，阻止了二十年的微流体研究提供引人注目的替代方案的工作马应用？蛋白质印迹和双向电泳（2DE）。蛋白质组学技术的突破是必不可少的：虽然基因组革命对理解生命过程产生了广泛的影响，但可以说更重要的是？蛋白质组学革命尚未实现。蛋白质与功能更直接相关，在生物化学上更复杂，数量比基因多10- 100倍。因此，几乎所有的生命科学研究都需要多步分析，如Western blot和2DE。然而，台式多步分离需要大量的劳动力，时间投资和珍贵的样品来产生半定量;这些分析将从集成中受益匪浅。独特的微马赛克集成策略-微室区域性地与离散的纳米结构分离材料-具有革命性的多步分离的潜力，以应对21世纪世纪紧迫的蛋白质组学挑战。多步蛋白质组学分析测量的不是一个，而是多个物理化学和功能特性。迄今为止，多步微量分离的性能受到将第一测定步骤（单个微通道）与作为微通道阵列的第二测定步骤耦合的现状设计策略的影响。遗憾的是这种做法？不可信吗第一级分离读数通过将它们映射到第二级中的离散隔室。为了克服集成损失，PI提出了一个从根本上脱离现状，通过引入微室与离散的纳米材料的区域性微图案。基础研究支撑的mosaic集成技术将包括：1）发展定量评估的微/纳米纤维技术和设备的电荷，化学和光图案化的聚合物，以实现本地化的生物化学/物理功能内的流体网络。图案化的纳米结构材料内的反应将使用荧光成像进行研究，确定特异性，动力学和总体亲和力，在对照实验中使用受控的均质试剂流。2)采用分散控制策略的分离集成，实现高性能多步分离。PI将研究电迁移，扩散和反应之间的相互作用经验，分析，并与三维多物理场求解器。PI将识别和控制传输和反应机制;即，空间非均匀表面反应的谱带展宽。虽然在均匀微通道网络中理解了耦合的分散和反应，但3D几何形状并非如此，3D非均相反应模式支撑了本文提出的微通道马赛克多步测定。总之，这些研究将提供新的知识：制造;纳米结构材料，分离设备和分析工程;实验方法。 虽然可用的台式蛋白质组学工具无处不在，但在通量、定量、动态范围和自动化方面存在显著的限制。拟议的新颖和广泛适用的工具将具体推进每个？影响系统生物学、合成生物学和生物燃料。可能的结果包括：基于模拟的治疗剂筛选、新型纤维素生物燃料的高通量鉴定以及细菌生物修复的合理选择算法。PI将通过两门正在开发的课程中的新多媒体讲座材料以及新的K-12合作伙伴关系（劳伦斯科学独创性实验室大厅）中的实践模块将研究与教学结合起来。PI提出了一个全面的战略，以招募和留住代表性不足的学生，并在三个NSF REU？S.作为伯克利的指导老师？的女工程师协会（SWE），她将支持SWE和伯克利？的新生研究生妇女在工程组通过NSF的支持。她通过会议主席、技术项目委员会服务和社区建设（例如，MEMS中的女性）。