Membrane Receptor Microarrays Based on Quantum Dot Barcoded Lipobeads

摘要提案标题：基于量子点条形码脂质微珠的膜受体微阵列，提案号：CTS-0626139，主要研究者：亚历山大Couzis，机构：纽约城市大学摘要：本项目将开发一种新的用于表面细胞膜受体显示的微阵列平台。该阵列预期用于高通量筛选膜受体与潜在结合配偶体的结合。在阵列设计中，受体被隔离在支持的双层中，其封装直径为1 m的条形码二氧化硅或聚苯乙烯珠以形成脂质珠。通过将脂质珠插入微米大小的威尔斯孔（比脂质珠稍大）中，将脂质珠排列在表面上。微威尔斯井在表面上以正方形网格图案化，其中威尔斯井由3 m的中心到中心间距分开。包封的颗粒使用由嵌入颗粒中的发光量子点制成的标记物进行条形码化，并共价连接至二氧化硅或聚苯乙烯基质。该阵列在1 mm x 1 mm的活动区域上有大约105个登记中心，其中每个微珠登记中心能够显示不同的受体。检测结合事件，并使用荧光显微镜读取识别脂质珠上受体的条形码。将通过筛选细胞膜隔离的糖脂与细菌毒素的相互作用来测试阵列。本文介绍了量子点编码珠的合成和微孔图案化表面的制造的初步结果。该计划的成功将对蛋白质的微阵列，特别是膜受体的微阵列产生重大影响：该平台允许小型化，其比用于展示膜受体或蛋白质的任何现有平台小几个数量级，并且提供了对脂质环境严格要求的难以展示的膜受体的容易展示。该阵列将提供的探测膜受体的结合相互作用的增强的能力对于药物发现是特别重要的，因为药物的分子靶标的50%是膜结合的。它也可以被证明是一个重要的工具，用于映射和理解细胞蛋白质组中多达25%的膜蛋白编码基因的结合相互作用。从更广泛的影响角度来看，该提案广泛涉及拟议工作的潜在经济和社会效益。 PI将参加CCNY的高中外展计划。该项目将包括研究生和本科生的培训将与PI合作，并通过CCNY现有的计划提供支持。PI已经证明，她可以很好地与本科生，这是一个宝贵的方面，拟议的工作。因此，该提案的教育方面是该提案的一个优势。

Abstract Proposal Title: Membrane Receptor Microarrays Based on Quantum Dot Barcoded Lipobeads, Proposal Number: CTS-0626139, Principal Investigator: Alexander Couzis, Institution: City University of New York Abstract:A new microarray platform for the display of surface cell membrane receptors is will be developed in this project. The array is intended to be used for the high throughput screening of the binding of membrane receptors with potential binding partners. In the array design, the receptors are sequestered in a supported bilayer, which encapsulates a 1 m in diameter barcoded silica or polystyrene bead to form a lipobead.Insertion of the membrane receptor in the bilayer allows the receptor to retain its biological activity. The lipobeads will be arrayed on a surface by inserting them intomicron sized wells (slightly larger than the lipobead). The micro-wells are patterned in asquare grid on the surface in which wells are separated by a center-to-center spacing of3m. The encapsulated particles are barcoded using a label made of luminescent quantumdots embedded into the particles, and covalently linked to the silica or polystyrenematrix. The array has approximately 105 registries on a 1mm x 1mm active area, with thelipobead registries capable of each displaying a different receptor. Binding events aredetected and the barcode identifying the receptor on the lipobead read using fluorescencemicroscopy. The array will be tested by screening the interaction of a cell membrane sequestered glycolipid with a bacterial toxin. Preliminary results are presented on the synthesis of the quantum dot encoded beads and the fabrication of the micro-well patterned surface.The success of this program will have a significant impact in the microarraying of proteins in general and membrane receptors in particular: The platform allows for a miniaturization that is orders of magnitude smaller than any current platforms for the display of membrane receptors or proteins in general, and provides for the facile display of difficult to exhibit membrane receptors with their strict requirement of a lipid environment. The enhanced ability to probe the binding interactions of membrane receptors which this array will afford is particularly important to drug discovery since fifty percent of the molecular targets of drugs are membrane bound. It could also prove be an important tool for the mapping and understanding of the binding interactions of the cell proteome in as much as twenty five percent of the sequenced genes of a given organism code for membrane proteins. From a broader impacts perspective, the proposal broadly addresses the potential economic and societal benefits of the proposed work. The PIs will participate in CCNY's high school outreach program. The project would include the training of a graduate student and undergraduates will work with the PI and be supported through pre-existing programs at CCNY. The PI has demonstrated that she can work well with undergraduate students, which is a valuable aspect of the proposed work. The education aspects of the proposal are thus a strength of the proposal.