Imaging supported lipid bilayers and actual cell membrane fragments by NanoSIMS

描述（由申请人提供）：支撑的脂质双层已被用作研究具有确定组成的模型膜的生物物理性质的工具。特别注意胆固醇对脂质膜相行为的作用，特别是脂质“筏”或结构域和复合物的形成。用于阐明二元和三元脂质双层（即荧光和原子力显微镜）的相行为的常见技术已被限制，其无法提供直接的信息的空间组成的膜。二次离子质谱（NanoSIMS）是一个强大的工具，成像的横向组织和组成的脂质双层。由于脂质双层中的组分由同位素标记唯一识别，因此NanoSIMS可以明确识别在小至50 nm的区域中共定位的不同分子。在这些研究中，描述了在高水平的13 C富集下生产胆固醇同位素异构体的方法。此外，NanoSIMS将用于成像天然细胞膜片段中胆固醇和蛋白质的空间分布和共定位。 公共卫生相关性：细胞膜的脂质双层在横向组成和组织方面是一个复杂的动态系统。活细胞依赖于膜组分的协同重排，如胆固醇，脂质和蛋白质;然而，很少有人知道这种组织在基本过程中如何变化，如细胞间相互作用，信号转导，膜运输和膜结合蛋白的活性和可及性。在这些研究中，二次离子质谱（nanoSIMS）成像将用于获得定量的组成信息与模型和实际细胞膜的纳米级分辨率。

DESCRIPTION (provided by applicant): Supported lipid bilayers have been used as a tool to study the biophysical properties of model membranes with defined compositions. Special attention has been given to the role of cholesterol on the phase behavior of lipid membranes, in particular, to the formation of lipid "rafts" or domains and complexes. Common techniques used to elucidate the phase behavior of binary and ternary lipid bilayers (i.e. fluorescence and atomic force microscopy) have been limited by their inability to provide direct information on the spatial composition of membranes. Secondary ion mass spectrometry (NanoSIMS) is a powerful tool for imaging the lateral organization and composition of lipid bilayers. Because the components in a lipid bilayer are uniquely identified by isotopic labels, NanoSIMS can unambiguously identify different molecules co-localized in a region as small as 50nm. In these studies, a method for producing cholesterol isotopomers at high levels of 13C enrichment is described. Furthermore, NanoSIMS will be used to image the spatial distribution and co-localization of cholesterol and proteins in native cell membrane fragments. PUBLIC HEALTH RELEVANCE: The lipid bilayer of a cell membrane is a complex and dynamic system in terms of lateral composition and organization. Living cells depend on the concerted rearrangement of membrane components, such as cholesterol, lipids, and proteins; yet very little is known about how this organization changes during essential processes, such as, cell-cell interactions, signal transduction, membrane trafficking, and membrane-bound protein activity and accessibility. In these studies, secondary ion mass spectrometry (nanoSIMS) imaging will be used to obtain quantitative compositional information with nanometer scale resolution of model and actual cell membranes.