Microfluidic MALDI-MS Device for High-Throughput Proteomics & Biomarker Discovery

描述(由申请人提供)：癌症是美国的主要死亡原因，每年有超过100万例新病例被诊断出来。由于大规模人群筛查所需的高敏感性和高特异性，目前在临床上常规使用的单一蛋白生物标记物很少。关键的优先事项是开发新的技术，使之能够快速检测与早期诊断、预后、分期和治疗反应相关的大量生物标志物。 这项研究的长期目标是将新兴的微流控技术与最先进的质谱学(MS)检测相结合，以提高我们分析生物系统中分子结构和功能的能力。该应用充分利用了微流控体系结构的独特能力，可实现微量样品的过程集成、多路复用、快速和高通量处理，以及MS检测的能力，可提供明确检测痕量组分所需的灵敏度、特异度和分辨率。 具体目标1：开发一种紧凑、低成本、一次性的微流控分析平台，用于高通量蛋白质组学的基质辅助激光解吸电离(MALDI)-MS/MS检测，这将使蛋白质共表达模式的研究和生物标志物的发现成为可能。该微流控设备将进行并行高效液相色谱(LC)分离，并将集成一种新型微芯片-MS接口，以实现直接从芯片进行灵敏的MALDI-MS/MS检测。 具体目标2：证明微流控MALDI-MS/MS平台在检测细胞提取物中多种癌症生物标志物方面的有效性。将对MCF7乳腺癌细胞系的细胞部分进行分析，以寻找参与推动癌症发生和发展的基本细胞过程(细胞增殖、细胞周期调节、DNA修复、细胞凋亡和侵袭/转移)的目标蛋白。

DESCRIPTION (provided by applicant): Cancer is a leading cause of death in the United States with over 1,000,000 new cases being diagnosed every year. As a result of the high sensitivities and specificities that are required to justify large-scale population screening, only very few single protein biomarkers are routinely used today in the clinical setting. It is of critical priority to develop novel technologies that will enable the rapid detection of a plethora of biomarkers relevant to early diagnosis, prognosis, staging and treatment response. The long-term objective of this research is to combine the emerging technology of microfluidics with state-of-the-art mass spectrometry (MS) detection to enhance our capacity for analyzing molecular structure and function in biological systems. This application capitalizes on the distinguishing capabilities of microfluidic architectures that enable process integration, multiplexing, fast and high-throughput processing of minute amounts of sample, and the power of MS detection that provides the sensitivity, specificity and resolving power necessary for unambiguous detection of trace level components. Specific Aim 1: Develop a compact, low-cost and disposable microfluidic analysis platform with matrix assisted laser desorption ionization (MALDI)-MS/MS detection for high-throughput proteomics that will enable the study of protein co-expression patterns and biomarker discovery. The microfluidic device will carry out parallel liquid chromatography (LC) separations and will integrate a novel microchip-MS interface to enable sensitive MALDI-MS/MS detection directly from the chip. Specific Aim 2: Demonstrate the effectiveness of the microfluidic MALDI-MS/MS platform for the detection of multiple cancer biomarkers in cellular extracts. Cellular fractions from the MCF7 breast cancer cell line will be analyzed for target proteins that are involved in essential cellular processes driving cancer on-set and development (cell proliferation, cell cycle regulation, DNA repair, apoptosis and invasion/metastasis).