Nanofluidic system for analysis of single biological molecules and particles

用于分析单个生物分子和颗粒的纳流体系统

• 批准号: 7572610
• 负责人: Rohit N Karnik
• 金额: $ 14.72万
• 依托单位: MASSACHUSETTS INSTITUTE OF TECHNOLOGY
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-02-01 至 2011-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7572610
• 关键词: Back; Binding; Biochemical; Biological; Biological Assay; Biological Models; Biology; Caliber; Cells; Characteristics; Charge; Clinical Research; Coulter counter; Cytometry; DNA; DNA Sequence; DNA Viruses; Detection; Development; Devices; Diagnostic; Digestion; Event; Feedback; Fingerprint; Foundations; Genomics; Goals; Healthcare; Hemolysin; Label; Laboratories; Length; Measurement; Microscopy; Miniaturization; Molecular; Nanosphere; Nanotechnology; Noise; Organelles; Physiologic pulse; Process; Proteins; Research; Resolution; Signal Transduction; Solutions; Surface; System; Systems Analysis; Techniques; Technology; Therapeutic; Time; Virus; Work; base; biological systems; design; gel electrophoresis; improved; lithography; nanochannel; nanofabrication; nanofluidic; nanoparticle; nanopore; nanoscale; new technology; particle; prevent; protein complex; public health relevance; sensor; single molecule; voltage

DESCRIPTION (provided by applicant): Technologies for analysis of biological molecules and particles are crucial for advancing research in biology and health care. There is a significant gap in technology for rapid, label-free characterization of unknown particles and molecules in the size range of 10-500 nm. While larger particles and cells are easily characterized by a variety of techniques including Coulter counters and cytometers, inadequate signal-to-noise ratio is a key limitation of detecting smaller particles using Coulter counters. Advances in nanotechnology have resulted in nanopore sensors based on the Coulter counter principle, which are being developed to a large extent for DNA sequencing. Their sensitivity arises due to their size being comparable to the size of the analyte. However, these sensors are typically not suitable for analysis of larger particles or molecules with varying unknown size, and their fabrication is also difficult. A common feature of nanopores and Coulter counters is that the analyte particle or molecule escapes into the solution after measurement, which limits the time during which the particle is analyzed and results in poor signal-to-noise ratio. However, if multiple measurements were possible on the same particle, the signal-to-noise ratio may be expected to be dramatically enhanced due to statistical averaging over the measurements. The goal of this project is to develop a nanofluidic Coulter counter system with feedback control that will enable multiple measurements on the same particle, thereby greatly improving the signal to noise ratio for sizing nanoscale analytes (Aim 1). This system will be evaluated for its ability to size fragments of genomic DNA (Aim 2) and to distinguish between nanospheres with slight differences in size as models for biological particles (Aim 3). This goal will be achieved using a system consisting of a nanochannel (50-500 nm) flanked by two nanofluidic reservoirs that serve to trap analytes and prevent their escape during measurement. The devices will be fabricated using soft lithography techniques that are amenable to integration into fluidic systems. The proposed device is the first step towards integrated systems capable of manipulation, processing, and analysis of single particles and molecules at the nanoscale. If successful, this project will lay the foundations for integrated fluidic systems for analysis of single particles, PCR-free assays, and may eventually enable simple biochemical analyses such as DNA and protein digestion and fingerprinting at the single molecule level. PUBLIC HEALTH RELEVANCE: Narrative In this project we will develop a nanofluidic system for sizing of large DNA fragments with a nanochannel sensor. This work may result in significant improvement in signal-to-noise ratios for label-free analysis of single biological particles and molecules in the nanoscale size range.

描述（由申请人提供）：生物分子和颗粒分析技术对于推进生物学和医疗保健研究至关重要。在对10-500 nm尺寸范围内的未知颗粒和分子进行快速、无标记表征的技术方面存在重大差距。虽然较大的颗粒和细胞很容易通过各种技术（包括库尔特计数器和细胞计数器）进行表征，但信噪比不足是使用库尔特计数器检测较小颗粒的关键限制。纳米技术的进步已经产生了基于库尔特计数器原理的纳米孔传感器，其在很大程度上被开发用于DNA测序。它们的灵敏度是由于它们的大小与分析物的大小相当。然而，这些传感器通常不适用于分析具有不同未知尺寸的较大颗粒或分子，并且它们的制造也很困难。纳米孔和库尔特计数器的一个共同特征是，分析物颗粒或分子在测量后逃逸到溶液中，这限制了分析颗粒的时间，并导致信噪比差。然而，如果对同一颗粒进行多次测量是可能的，则由于对测量进行统计平均，可以预期信噪比会显著增强。该项目的目标是开发一种具有反馈控制的纳米流体库尔特计数器系统，该系统将能够对同一颗粒进行多次测量，从而大大提高测量纳米级分析物的信噪比（目标1）。将评价该系统测定基因组DNA片段大小的能力（目标2），以及区分大小略有差异的纳米球作为生物颗粒模型的能力（目标3）。这一目标将使用由两侧为两个纳米流体储库的纳米通道（50-500 nm）组成的系统来实现，所述纳米流体储库用于捕获分析物并防止它们在测量期间逃逸。这些器件将使用软光刻技术制造，这些技术可以集成到流体系统中。该设备是迈向集成系统的第一步，该系统能够在纳米尺度上操纵、处理和分析单个粒子和分子。如果成功，该项目将为用于分析单个颗粒、无PCR检测的集成流体系统奠定基础，并可能最终实现简单的生化分析，如DNA和蛋白质消化以及单分子水平的指纹分析。 公共卫生相关性：叙述在这个项目中，我们将开发一个纳米流体系统的大小与纳米通道传感器的大DNA片段。这项工作可能会导致显着改善的信号-噪声比的无标记分析的单个生物颗粒和分子在纳米尺寸范围内。