Collaboration in the Development of Novel Polymerization Enhanced Immunofluoresce

描述(申请人提供)：本次合作将开发新一代成像技术，广泛应用于生物和医学的所有领域。这个项目旨在应用许多众所周知的聚合原理来开发一种新的免疫荧光标记策略。我们的方法被称为基于聚合的扩增(PBA)，它基于获得聚合所特有的特异性和高度放大的响应，以响应通过生物识别事件耦合到感兴趣的细胞或亚细胞特征的起始分子。具体地说，我们将开发一种方法，利用聚合起始分子与抗体的偶联，基于蛋白质表达在局部区域产生聚合。我们定制了~100 nm的聚合物薄膜，通过在聚合物网络中捕获荧光纳米颗粒来包含大量的荧光标签。初步结果表明，使用我们的PBA技术，一次抗体使用量可以减少三个数量级以上。标记的基于包埋法的固定化使得能够使用许多商业上可获得的荧光团，而不需要对荧光纳米颗粒进行化学修饰。我们寻求开发这项技术，以解决跨越生物学和临床研究的问题。为此，我们将开发PBA系统，以满足培养细胞、多目标/多颜色、组织和基于流程的应用。这项技术的发展将使利用廉价的成像仪器检测低丰度蛋白质成为可能。此外，抗体消耗量减少三个数量级将大幅减少基础生物和临床研究的资金需求。这种成本节约将不仅转化为个人研究实验室，而且还将转化为研究资助机构。这个应用程序建立了适合项目的现有调查人员配对，每个人都在各自的领域拥有持续的成就记录。鲍曼博士是光聚合反应工程方面的专家，在开发和表征该项目所需的先进聚合物系统方面拥有丰富的物理资源。Balasubramaniam博士是儿科肺病学的积极研究人员，并带来了免疫荧光技术和组织学方面的专业知识。Balasubramaniam博士是科罗拉多大学丹佛健康科学中心儿科心肺实验室的主任，可以访问此次合作所需的所有生物学特征资源。 与公共卫生相关：该项目协同结合使用聚合物化学和生物学，以促进检测存在的数量比以前可能少得多的生物物种。检测少量生物物种的能力将使生物学研究人员能够以低廉的成本探索细胞和组织的新科学方面。

DESCRIPTION (provided by applicant): This collaboration will develop a next generation imaging technology for broad application across all fields of biology and medicine. This project aims to apply many well understood principles of polymerization to the development of a new strategy for immunofluorescent labeling. Our approach, termed polymerization- based amplification (PBA), is based on obtaining a specific and highly amplified response unique to polymerization in response to an initiating molecule that is coupled through a biorecognition event to the cell or sub-cellular feature of interest. Specifically, we will develop the approach to utilize the coupling of a polymerization initiating molecule to an antibody to generate polymerization in localized regions, based on protein expression. We tailor the ~100 nm polymer film to incorporate numerous fluorescent labels through entrapment of fluorescent nanoparticles in the polymer network. Preliminary findings demonstrate over a three order of magnitude reduction in primary antibody utilization is achievable with our PBA technology. The entrapment based immobilization of the label enables the use of numerous commercially available fluorophores, without the need for chemical modification of the fluorescent nanoparticle. We seek to develop this technology to address problems spanning biology and clinical research. To that end, we will develop PBA systems to address cultured cell, multi-target / multi-color, tissue, and flow-based applications. The development of this technology will enable the detection of low abundance proteins with inexpensive imaging instrumentation. In addition, a three order of magnitude reduction in antibody consumption would provide a drastic decrease in financial requirements for basic biological and clinical research. This cost savings would translate not only to the individual research lab, but also to research funding agencies. This application establishes a project-appropriate pairing of established investigators, each with an ongoing record of accomplishments in their respective field. Dr. Bowman is an expert in photopolymerization reaction engineering, and has extensive physical resources for the development and characterization of the advanced polymer systems required by this project. Dr. Balasubramaniam is an active researcher in pediatric pulmonology and brings expertise in immunofluorescent techniques and histology. Dr. Balasubramaniam is the Director of the Pediatric Heart Lung Lab at the University of Colorado - Denver Health Sciences Center, and has access to all of the biological characterization resources required by this collaboration. PUBLIC HEALTH RELEVANCE: This project synergistically combines uses polymer chemistry and biology to facilitate the detection of biological species that are present in amounts much lower than previously possible. The ability to detect small amounts of biological species will allow researchers in biology to inexpensively explore new scientific aspects of cells and tissues.