Cancer Sample Preparation with Micromachined Magnetic Sifter and Nanoparticles

描述（由申请人提供）：样品制备的差异可能导致不同实验室鉴定的生物分子或细胞的数量和类型存在重大差异，即使使用相同的试剂和生物传感器（或生物芯片）。这尤其阻碍了我们对抗癌症的努力，因为样品制备是癌症生物标志物发现、验证及其在患者分子诊断和治疗中的应用的先决条件。这种对癌症样品制备的迫切需求需要创新的方法来带来更好和更实惠的样品制备工具。我们已经组建了一个跨学科的团队，拥有来自磁学、纳米技术、癌症临床实践和研究、生物化学和蛋白质组学的专业知识，致力于新型微机械磁性筛和直接制造磁性纳米颗粒。磁性筛可用于分离和富集癌症靶点；包括原始样本中的蛋白质和稀有细胞。精确尺寸结构的利用，包括亚纳米控制量子磁效应的使用，将实现高通量、高捕获收率、低杂质和低成本。该技术非常独特，因为通过三维硅基微加工可以实现高磁场梯度（>特斯拉/ 5m）和大流速（> 1ml /小时）。使用具有独特形状和可控磁链形成的高力矩磁性纳米颗粒，可以提高前所未有的捕获和释放效率，最大限度地减少细胞损伤，增强表征和运行到运行的可重复性。拟议的项目有三个目标：具体目标1。高效捕获和分析蛋白质癌变标志物和癌细胞的磁筛：a)构建高磁场梯度（>1 Tesla/ 5m）、大流速（>1 mL/h）的磁筛，演示高效捕获、释放和分析蛋白质癌变标志物。b)利用磁筛捕获并释放NCI-H1650肺癌肿瘤细胞，将癌细胞加入到人血样品中，总体捕获效率为0.70%，同时保持细胞存活。具体目标2。用于高效高速磁分离的新型磁性纳米颗粒的功能化与表征。具体目标3。磁性筛用于从人体血液中捕获和分析稀有细胞。这笔资金的用途包括大幅减少分离后分析，并在目前难以获得和分析的稀有分子和细胞类型领域开展新的研究。此外，将蛋白质和癌症标志物从血液中的高浓度杂质中分离出来，富集1000倍，将能够检测到传统技术难以量化的低丰度癌症标志物。我们期望在这项资助下开发的工具将对癌症研究和治疗界产生重大影响。

DESCRIPTION (provided by applicant): Variations in sample preparation may contribute to major discrepancies in the quantity and type of biomolecules or cells identified by different laboratories, even though the same reagents and biosensors (or biochips) are employed. This is especially hampering our efforts to fight cancer as sample preparation is a prerequisite for cancer biomarker discovery, validation, and their use in molecular diagnosis and treatment of patients. This urgent need for cancer sample preparation requires innovative methods to bring about better and more affordable sample preparation tools. We have formed an interdisciplinary team with expertise from magnetics, nanotechnology, cancer clinical practice and research, biochemistry, and proteomics to work on a novel micromachined magnetic sifter and directly fabricated magnetic nanoparticles. The magnetic sifter can be used to separate and enrich cancer targets; including proteins and rare cells from raw samples. The utilization of precisely dimensioned structures, including the use of sub-nanometer control of quantum magnetic effects will enable high throughput, high capture yield, low impurities, and low cost. The technology is very unique because high magnetic field gradients (>1 Tesla/ 5m) and large flow rates (>1 mL/hour) are enabled by three-dimensional Si-based micromachining. The use of high-moment magnetic nanoparticles with distinctive shapes and controlled magnetic chain formation give rise to unprecedented capture and release efficiencies, cell damage minimization, enhanced characterization, and run-to-run reproducibility. The proposed project is organized in three aims: Specific Aim 1. Magnetic sifter for high efficiency capture and analysis of protein cancer markers and cancer cells: a) Construct a magnetic sifter with a high magnetic field gradient (>1 Tesla/ 5m) and large flow rate (>1 mL/hour) and demonstrate the efficient capture, release and analysis of protein cancer markers. b) Use magnetic sifter to capture and release NCI-H1650 lung cancer tumor cells cancer cells spiked into samples of human blood with an overall capture efficiency of >70%, while keeping cells viable. Specific Aim 2. Functionalization and characterization of novel magnetic nanoparticles for high efficiency high speed magnetic separation. Specific Aim 3. Magnetic sifter for rare cells capture and analysis from human blood. The utility of the grant includes drastically reducing post-separation analysis and enabling new investigations in areas where rare molecules and cell types are currently too difficult to obtain and analyze. Furthermore, separating protein and cancer markers from high concentration impurities in blood, with 1000-fold enrichment, will enable detection of low abundance cancer markers which are difficult to quantify with conventional technologies. We expect that the tools developed under this grant will have a great impact on the cancer research and treatment community.