Observing the IgE and FceRI signaling pathway via dielectric-filled nanoapertures

描述（由申请人提供）：本研究旨在了解过敏原激发免疫系统反应的分子过程。特别是，我们打算推进理解的细胞内过程，导致抗体诱导的膜受体的交联，并诱导过敏和炎症反应。该项目将通过开发一系列孔径来提高超分辨率显微镜的实验能力，每个孔径利用近场光学照亮一个小的样品体积。 这项研究的长期目标是了解用于修改质膜局部组成的物理和化学过程。本项目的目的是测量膜重组和信号传导过程与nano-patterened光学表面能够80 nm和1 ps的分辨率，否则传统的荧光显微镜技术。该项目将产生一种新技术，以克服目前的实验障碍，并应用于各种生物学假设。本研究的短期目标是（1）开发一种能够在浓缩生物环境中进行单分子检测的技术，以及（2）进一步了解IgE-FccRI聚类、蛋白募集和局部膜异质性的诱导。这项研究的中心假设是，介质填充的零模式波导将提供足够的时间和空间分辨率的细胞膜上测量蛋白质-蛋白质协会由受体刺激诱导。这一假设建立在相关的纳米结构和理论预测的基础上，如下所述。这种装置将提供一种测量膜（重组）的手段，测试以前无法回答的假设，并推进基于膜受体的营养物的治疗。这个项目提供了一个独特的培训机会，以建立在我的生物物理化学背景，在纳米纤维和膜生物学指导。 这项研究将创造一种新的技术，并应用它来回答紧迫的问题，在细胞内信号级联后抗原刺激和交联的FC？RI膜受体。这项研究的结果将直接适用于诊断和治疗过敏反应。具体目标1：开发近场光学显微镜（ANOMs）的孔径阵列具体目标2：表征定量超分辨率显微镜的ANOMs的光学特性具体目标3：测量FC？RI聚类

DESCRIPTION (provided by applicant): This research aims to understand the molecular processes by which an allergen elicits an immune system response. In particular, we intend to advance understandings of the intracellular processes that result from antibody-induced cross-linking of the membrane receptors and induce an allergic and inflammation response. This project will advance the experimental capabilities for super-resolution microscopy through the development of an array of apertures, each illuminating a small sample volume utilizing near-field optics. The long-term goal of this research is to understand the physical and chemical processes used to modify local composition of the plasma membrane. The objective of this project is to measure the membrane reorganization and signaling processes with nano-patterened optical surfaces capable of 80 nm and 1 ps resolution with otherwise conventional fluorescent microscopy techniques. This project will yield a new technology to overcome current experimental barriers and have applications to a variety of biological hypotheses. The short-term goals of this study are (1) to develop a technique capable of single-molecule detection in concentrated biological environments and (2) to further understand IgE-FccRI clustering, protein recruitment, and the induction of local membrane heterogeneity. The central hypothesis of this research is that dielectric-filled zero-mode waveguides will provide sufficient time and space resolution on cell membranes to measure protein-protein associations induced by receptor stimulation. This hypothesis is built upon both related nanoscale structures and theoretical predictions, as described below. Such devices will provide a means for measuring membrane (re)organization, testing previously unanswerable hypotheses, and advancing the treatment of membrane-receptor based aliments. This project provides a unique training opportunity to build upon my background in biophysical chemistry, guided by mentoring in nanofabrication and membrane biology. This research will create a new technology and apply it to answer pressing questions in the intracellular signaling cascade following antigen stimulation and cross-linking of FC?RI membrane receptors. Results from this research will be directly applicable to diagnosing and treating allergic responses. Specific Aim 1: Develop an array of apertures for near-field optical microscopy (ANOMs) Specific Aim 2: Characterize the optical properties of ANOMs for quantitative super-resolution microscopy Specific Aim 3: Measure local membrane heterogeneity induced by FC?RI clustering