RIG: Metabolite Target Analysis using Microdevice Electrophoresis

该项目将开发新的微设备(微流控)电泳法，以改进某些细胞过程特有的代谢物的分离和检测。电泳是一种基于电场中不同的迁移率来分离分子的技术，它仍然是生物学和化学中最强大和最广泛使用的分析工具之一。与传统的平板凝胶相比，现代毛细管电泳法具有许多优点，如灵敏的在线检测、速度快、成本低、焦耳散热更好。将使用两种不同的策略创建分析雌激素代谢物的微流控方法，即自由溶液和水凝胶固定化。这两种方法都将利用竞争性免疫分析。自由溶液微流体法将使用传统的竞争免疫分析方法，而水凝胶固定化策略将依赖于在丙烯酰胺凝胶塞中物理捕获针对雌激素代谢物的抗体。将评估抗原特异性水凝胶捕获和浓缩不同浓度雌激素代谢物的能力。将评估检测限和非特异性结合，以直接比较这两种方法的实用性和不同的应用。这项研究的更广泛的影响与它的潜力有关：1.在跨学科研究方面培训众多少数族裔研究生和博士后；2.加强当前的外展活动，旨在为经济和社会上处于不利地位的高中生提供实践研究经验；3.与微流控电泳法相关的先锋知识；以及4.彻底改变了科学家分析大分子的速度和分辨率。

This project will develop new microdevice (microfluidic) electrophoresis methods to improve the separation and detection of metabolites unique to certain cellular processes. Electrophoresis, a technique used to separate molecules based on differential migration rates in an electric field, remains one of the most powerful and widely used analytical tools in biology and in chemistry. The modern capillary format, microdevice electrophoresis, offers numerous advantages over the traditional slab gel, such as sensitive on-line detection, speed, cost, and improved dissipation of joule heating. Microfluidic methods to analyze estrogen metabolites will be created using two different strategies, free solution and hydrogel immobilization. Both approaches will utilize competitive immunoassays. The free solution microfluidic method will employ conventional competitive immunoassays whereas the hydrogel immobilization strategy will rely on the physical entrapment of antibodies specific for estrogen metabolites within acrylamide gel plugs. Antigen specific hydrogels will be evaluated for their ability to capture and concentrate estrogen metabolites at various concentrations. Limits of detection and non-specific binding will be assessed to directly compare the utility and diverse applications of the two methods. The broader impacts of this research are linked to its potential to:1. train numerous minority graduate and postdoctoral students in interdisciplinary research, 2. enhance current outreach activities designed to give economically and socially disadvantaged high school students hands-on research experience, 3. pioneer knowledge relating to microfluidic electrophoresis, and4. revolutionize the speed and resolution with which scientists can analyze macromolecules.