Phase-Changing Sacrificial Layer Microfluidics for Enhanced Protein Analysis

不断开发新的工具和技术，以提高灵敏度、选择性、速度和 生物分析中的通量在降低每次测定的成本的同时，在临床诊断中将是关键的， 医学研究和生命科学的其他学科。微机械加工方法具有潜力， 能够构建低成本但复杂的微流体系统，用于分析复杂蛋白质 混合物。在此，我们提出发展一种相变牺牲层（PCSL）微细加工技术 方法和聚合物表面改性方法来制造廉价的微流体装置， 改善甲胎蛋白（AFP）的检测和定量，这是一种癌症和胎儿健康的标志物， 血液和尿液这项建议有四个目标。首先，我们将推广PCSL溶剂 键合制造技术和原子转移自由基聚合表面改性方法， 一系列用于蛋白质分析的聚合物微流体系统。其次，我们计划评估PCSL溶剂 用于制造复杂的多层微流体阵列的结合方法 加工步骤。第三，我们将开发这些制造技术， 免疫亲和纯化系统和蛋白质样品预浓缩器 微芯片毛细管电泳第四，我们将利用前三个目标中的工具， 从生物样品中提取和预浓缩低丰度蛋白质（如AFP）的微型设备 混合物;然后我们将评估这些微系统用于测定血液和尿液中的AFP水平。 重要的是，虽然该提案中的研究是针对开发用于检测肿瘤的小型化测定法， AFP，这些相同的制造和分析过程可以以简单的方式概括为 允许快速定量其他低丰度癌症和疾病生物标志物。因此，PCSL 技术和聚合物表面衍生化方法应该提供广泛适用的微芯片 用于电泳分析和定量人体蛋白质或其他生物分子的平台。

The continued development of new tools and techniques that improve sensitivity, selectivity, speed and throughput in biological analysis, while reducing the cost per assay, will be critical in clinical diagnosis, medical research, and other disciplines in the life sciences. Micromachining methods have the potential to enable the construction of low-cost, yet sophisticated microfluidic systems for the analysis of complex protein mixtures. Herein, we propose to develop a phase-changing sacrificial layer (PCSL) microfabrication approach and polymer surface modification methods to create inexpensive microfluidic devices that can improve the detection and quantification of alpha-fetoprotein (AFP), a cancer and fetal wellness marker, in blood and urine. The objectives of this proposal are four-fold. First, we will generalize the PCSL solvent bonding fabrication technique and atom-transfer radical polymerization surface modification methods to a range of polymer microfluidic systems for protein analysis. Second, we plan to evaluate the PCSL solvent bonding approach for making complex, multilayer microfluidic arrays that facilitate the integration of sample processing steps. Third, we will develop these fabrication techniques to enable the incorporation of immunoaffinity purification systems and protein sample preconcentrators for use in enhancing detection in microchip capillary electrophoresis. Fourth, we will utilize the tools from the first three objectives to make microdevices that extract and preconcentrate lower-abundance proteins such as AFP from biological mixtures; we will then evaluate these microsystems for the determination of AFP levels in blood and urine. Importantly, while the studies in this proposal are directed toward the development of miniaturized assays for AFP, these same fabrication and analysis procedures can be generalized in a straightforward manner to allow the rapid quantification of other lower-abundance cancer and disease biomarkers. Thus, PCSL techniques and polymer surface derivatization methods should provide a broadly applicable microchip platform for the electrophoretic analysis and quantification of proteins or other biomolecules in humans.