Collaboration in the Development of Novel Polymerization Enhanced Immunofluoresce

描述（由申请人提供）：这项合作将开发下一代成像技术，广泛应用于生物学和医学的所有领域。本项目旨在将许多已被充分理解的聚合原理应用于免疫荧光标记的新策略的开发。我们的方法，称为基于聚合的扩增（PBA），是基于响应于通过生物识别事件偶联到感兴趣的细胞或亚细胞特征的起始分子而获得聚合所特有的特异性和高度扩增的响应。具体而言，我们将开发的方法，利用聚合引发分子的抗体的耦合，以产生聚合在局部区域，基于蛋白质的表达。我们定制了~100 nm的聚合物膜，通过在聚合物网络中捕获荧光纳米颗粒来掺入许多荧光标记。初步研究结果表明，使用我们的PBA技术，一抗利用率可降低三个数量级以上。标记的基于包埋的固定使得能够使用许多可商购的荧光团，而不需要荧光纳米颗粒的化学修饰。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，以解决培养细胞，多目标/多色，组织和基于流动的应用。这项技术的发展将使低丰度蛋白质的检测与廉价的成像仪器。此外，抗体消耗量减少三个数量级将大大减少基础生物学和临床研究的资金需求。这种成本节约不仅会转化为单个研究实验室，而且会转化为研究资助机构。 该应用程序建立了一个适合项目的常设调查员配对，每个人都有在各自领域取得成就的持续记录。Bowman博士是光聚合反应工程方面的专家，拥有丰富的物理资源，可用于开发和表征该项目所需的先进聚合物系统。Balasubramaniam博士是一位活跃的儿科肺病研究人员，并在免疫荧光技术和组织学方面拥有专业知识。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表征资源。 公共卫生相关性：该项目协同结合使用聚合物化学和生物学，以促进检测生物物种，这些生物物种的存在量远远低于以前的可能性。检测少量生物物种的能力将使生物学研究人员能够廉价地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.