Experimental and Computational Design of a Microfluidic Device for Micro-Barcode Based Oligonucleotide Synthesis

美国国家科学基金会化学与运输系统分部？微粒和多相过程项目（1415）提案号：0729771主要研究者：Shaqfeh， Eric隶属：斯坦福大学提案题目：基于微条形码的寡核苷酸合成微流体装置的实验和计算设计微条形码技术有可能在单个反应容器中实现数十万个生化反应的大规模多路传输。微阵列在二维底物上进行大规模多路复用，通过对遗传变异和功能进行全基因组范围的研究，已经改变了生物医学研究。微型条形码粒子准备将这种能力转化为三维，自由解决格式，极大地扩展了这种强大技术的可能性。直径为0.25至1微米，长度为2至10微米的棒状金属颗粒（Nanoplex, Menlo Park， CA）可以与金属条纹一起生长，以10位的顺序编码信息。每个微型条形码颗粒携带一个可识别的特征，类似于传统的条形码，作为一种机制，跟踪分子探针，如寡核苷酸，附着在颗粒表面。然后将许多这样的颗粒混合，与样品反应，并在单个腔室或流体通道中平行检测。我们建议开发一种适用于大规模平行寡核苷酸合成的自动粒子流控制、电场对准、分类和读出技术。我们将建立和演示定制设计的微流体装置，这是第一次，将控制，读取和分类微流体系统中的微条形码颗粒。对设备设计和优化至关重要的是，将发展广义电动力学模型，该模型将考虑粒子布朗运动、电泳（包括单极和多极电动力学效应）、水动力和受限几何中的沉积。这些模型将采用大规模多粒子模拟的形式，使用与实验共同开发的新型数值代码。大规模的模拟将使我们能够准确地预测粒子在设备中的位置、速度和方向，从而定量地预测设备的性能。我们已经进行了初步的实验，在直流和交流电场的沉降条件下，我们对准并随后跟踪长5微米，直径0.25微米的圆柱形粒子的位置和方向。除了最初的实验之外，我们已经开发了模拟工具来模拟大量布朗棒在低雷诺数下的电泳排列和周期性系统中粒子诱导的电泳流的沉积，从而确定我们最终将详细研究的初始流动参数制度。我们工作的更广泛的研究影响和智力价值包括对悬浮力学中许多未解决的问题的基本理解，这些问题直接影响这些条形码读取器的性能。这些问题包括发展我们对以下方面的理解：(a)棒状聚合物和棒状胶体颗粒悬浮液从稀到半稀的流变性，包括ICEP相互作用；(b) ICEP流动对与纤维悬浮液同时沉降和平均流动有关的集体现象的影响；(c)剪切诱导扩散对棒状颗粒质心运动的影响；(d)非局部流动中，即平均流动尺度在杆长数量级的流动中，杆悬架的集体动力学。事实上，尽管这些原理是复杂流体的微流体学的内在原理，但这些非线性物理的许多组合将首次得到检验。此外，通过与斯坦福大学科学教师暑期研究计划的合作，将为高中科学教师提供两个暑期实习机会，以开发与微流控装置设计中使用大规模计算相关的广泛教育影响。这些实习将包括一到两所大学的教员。一个实习将与研究的实验方面有关，另一个与计算设计方面有关。实习将允许教师与pi和研究生密切合作，并组成一个工作组，了解微流控技术和先进计算作为工程设计工具的应用，然后将实验专业知识，演示和计算机模拟带回课堂。

National Science Foundation - Division of Chemical &Transport Systems ? Particulate & Multiphase Processes Program (1415)Proposal Number: 0729771 Principal Investigators: Shaqfeh, Eric Affiliation: Stanford Proposal Title: Experimental and Computational Design of a Microfluidic Device forMicro-Barcode Based Oligonucleotide Synthesis Micro-barcode technologies have the potential to realize large-scale multiplexing of hundreds of thousands of biochemical reactions in a single reaction vessel. Microarrays, which perform large-scale multiplexing on two-dimensional substrates, have transformed biomedical research by enabling genome wide investigation of genetic variation and function. Micro-barcode particles are poised to translate this capability to a three-dimensional, free-solution format, greatly expanding the possibilities of this powerful technology. Rod-shaped metallic particles with 0.25 to 1 micron diameters and lengths of 2 to 10 microns (Nanoplex, Menlo Park, CA) can be grown with metallic stripes that encode on the order of 10 bits of information. Each micro-barcode particle carries an identifiable signature, analogous to a conventional barcode, that serves as a mechanism for tracking molecular probes, such as oligonucleotides, attached to the particle surface. Many such particles then be mixed, reacted with a sample, and detected in parallel in a single chamber or fluidic channel. We propose to develop an automated particle flow control, electricfield alignment, sorting, and readout technology applicable to massively-parallel oligonucleotide synthesis. We will build and demonstrate custom-designed microfluidic devices, that for the first time, will control, read, and sort micro-barcode particles in microfluidic systems. Critical to the device design and optimization, will be the development of generalized electro-kinetic models that account for particle Brownian motion, electrophoresis (including mono- and multi-pole electrokinetic effects), hydrodynamic forces, and sedimentation in confined geometries. These models will be in the form of large-scale multi-particle simulations using novel numerical codes being developed jointly with the experiments. The large-scale simulations will allow us to accurately predict the location, velocity and orientation of the particles as they travel through the device, and thus quantitatively predict device performance.We have performed preliminary experiments in which we align and subsequently track the positions and orientations of cylindrical particles 5 microns long and 0.25 microns in diameter under settling conditions in both DC and AC electric fields. In additional to the initial experiments, we have already developed simulation tools to model the sedimentation of a large number of Brownian rods at low Reynolds number with electrophoretic alignment and particle-induced electrophoretic flow in periodic systems, thus determining the initial flow parameter regimes that we will ultimately examine in detail. The broader research impact and intellectual merit of our work includes a fundamental understanding of a number of unsolved problems in suspension mechanics which directly bear on the performance of these barcode readers. These issues include developing our understanding of (a) the rheology of rod-like polymer and rod-like colloidal particle suspensions from dilute through semi-dilute including ICEP interactions; (b) the effect of ICEP flow on the collective phenomena associated with the simultaneous sedimentation and mean flow of fiber suspensions; (c) the action of shear-induced diffusion on the center of mass motion of the rod-like particles; and (d) the collective dynamics of rod suspensions in non-local flows, i.e. those in which the mean flow scale is on the order of the length of the rod. Indeed, even though these principles are intrinsic to the science of the microfluidics of complex fluids, many of the combinations of these nonlinear physics will be examined for the first time. Moreover, the broad educational impact associated with using large scale computing for design of microfluidic devices will be developed as an integral part of two summer internships for high school science teachers via a partnership with Stanford's Summer Research Program for Science Teachers. These internships will include faculty at one or two Title I schools. One internship will be associated with the experimental aspects of the research and the other with the computational design aspects. The internships will allow the faculty members to work closely with the PIs and graduate students and form a working group to understand the applications of microfluidic technology and advanced computing as an engineering design tool, and thereafter take experimental expertise, demonstrations and computer simulations back to the classroom.