CAREER: IDBR: Ultrasmooth Patterned Metals for Membrane Biology

细胞膜是由不同的脂类组成的复杂的马赛克，胆固醇、鞘脂和膜蛋白形成了称为脂筏的纳米尺度区域。脂筏瞬间分隔膜成分，进而调节细胞在空间和时间上的功能。为了定量研究脂筏和整合在脂筏中的蛋白质的形成、形态和信号转导机制，必须在与真实细胞膜非常相似的人工纳米环境中进行探索。因此，将脂膜与能够更好地模拟自然细胞环境的工程固态传感器相结合的分析仪器将提供对重要的体内反应和过程的更深层次的洞察。拟议的平台将把柔软的脂类物质与纳米多孔金属膜的天然传感能力相结合。也就是说，表面等离子体共振(SPR)和电化学传感将被用来探索由脂筏和膜蛋白介导的生物过程。然而，SPR所需的金或银等贵金属通常不适用于纳米级的大面积、低成本图案化。为了克服这一挑战，模板剥离方法将被用于可重复地高通量制造超光滑纳米多孔金属薄膜，该薄膜将被用作集纳米流体、光学检测、光谱和电化学传感于一体的多功能平台。能够进行分子结合和转运分析的膜生物传感平台的开发将为探索细胞膜的动态异质性及其实现的基本生命过程提供新的工具。总体而言，这些仪器的传播将使生物学家能够使用精确设计的仿生纳米环境来控制、成像和定量分析脂膜及其成分，潜在地深入到单分子水平。为了更广泛地推广，PI将在每年4月为期一周的NanoDays活动期间，在明尼苏达州科学博物馆组织“与科学家坐在一起”会议，并建立一个互动活动站。该馆的展览摊位将以“宏观至微流体”为主题，为K-12学生现场演示微流体设备。与该项目相关的研究生和本科生将获得跨学科培训，范围从纳米制造和光学到化学和细胞膜。妇女和代表性不足的少数民族将有机会通过NSF REU计划体验纳米制造。PI关于生物仪器的新研究生课程将通过理论讲座和每周的动手实验室会议来整合他的研究和教育，教授膜生物传感器的构造和操作。新的仪器和技术将通过一年两次、为期两天的生物微机械系统短期课程向研究界传播。将演示如何制作光滑的图案化金膜、与微流体集成、脂膜形成、随后的SPR和电化学传感，使其他研究人员能够复制芯片制造和仪器组装。

Cell membranes are a complex mosaic of different lipids, with cholesterol, sphingolipids and membrane proteins forming nanoscale domains called lipid rafts. Lipid rafts transiently compartmentalize membrane constituents, which in turn mediate cellular functions in space and time. In order to quantitatively investigate the formation, morphology and signal transduction mechanisms mediated by lipid rafts and proteins integrated therein, they must be probed in artificial nano-environments that closely resemble real cell membranes. Analytical instruments that integrate lipid membranes with engineered solid-state sensors that can better mimic natural cellular environments will therefore offer deeper insights into important in vivo reactions and processes.The proposed platform will integrate soft lipid matter with the native sensing capability of nanoporous metallic films. Namely, surface plasmon resonance (SPR) and electrochemical sensing will be used to probe the biological processes mediated by lipid rafts and membrane proteins. However, noble metals such as gold or silver required for SPR have typically not been amenable to large-area, low-cost patterning at nanometer-scale resolution. To overcome this challenge, template-stripping methods will be used for reproducible high-throughput fabrication of ultrasmooth nanoporous metallic films that will be used as a multi-functional platform combining nanofluidics, optical detection, spectroscopy, and electrochemical sensing. The development of a membrane biosensing platform capable of molecular binding and transport assays will provide new tools to probe the dynamic heterogeneity of cellular membranes and the fundamental life processes they enable. Overall, disseminating these instruments will enable biologists to control, image, and quantitatively analyze lipid membranes and their constituents, potentially down to the single-molecule level, using precisely engineered biomimetic nano-environments.For broader outreach, the PI will organize "Sit with a Scientist" sessions and build an interactive Activity Station at the Science Museum of Minnesota during the one week-long NanoDays event in April of each year. The PI's exhibition booth at the museum will feature the theme of "Macro- to Microfluidics", with on-site demos of microfluidic devices for K-12 students. Graduate and undergraduate students associated with the project will gain interdisciplinary training ranging from nanofabrication and optics to chemistry and cellular membranes. Women and underrepresented minorities will be given opportunities to experience nanofabrication through NSF REU programs. The PI's new graduate course on biological instrumentation will integrate his research and education through theoretical lectures and weekly hands-on lab sessions to teach the construction and operation of membrane biosensors. New instruments and technology will be disseminated to the research community through an already active biannual, two-day bioMEMs short course. The process of making smooth patterned gold films, integration with microfluidics, lipid membrane formation, followed by SPR and electrochemical sensing will be demonstrated, allowing other researchers to duplicate the chip fabrication and instrument buildup.