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

国家科学基金会--化学与运输系统部门？粒子和安培多阶段过程计划(1415年)建议编号：0729771主要研究人员：Shaqfeh，Eric隶属关系：斯坦福大学建议标题：基于微条码的寡核苷酸合成微流控设备的实验和计算设计微条码技术有可能在单个反应容器中实现数十万种生化反应的大规模多路复用。微阵列在二维基质上进行大规模的多路复用，通过使全基因组研究遗传变异和功能成为可能，改变了生物医学研究。微型条形码颗粒准备将这一能力转化为三维、自由溶液格式，极大地扩展了这项强大技术的可能性。直径0.25到1微米、长度2到10微米的棒状金属颗粒(Nanopex，Menlo Park，CA)可以用编码约10比特信息的金属条带生长。每个微型条形码颗粒都带有一个可识别的签名，类似于传统的条形码，用作跟踪附着在颗粒表面的分子探针(如寡核苷酸)的机制。然后，许多这样的颗粒被混合，与样品反应，并在单个腔室或流体通道中并行检测。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、分选和读出技术。我们将建造和演示定制设计的微流控设备，这将首次控制、读取和分选微流控系统中的微条形码颗粒。对设备设计和优化至关重要的是发展广义电动模型，该模型解释了颗粒布朗运动、电泳(包括单极和多极电动效应)、流体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，使用与实验联合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通过器件时的位置、速度和方向，从而定量地预测器件的性能。我们已经进行了初步的实验，在直流和交流电场中的稳定条件下，我们对准并随后跟踪了长5微米和直径0.25微米的圆柱形粒子的位置和方向。除了最初的实验，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沉积和周期性系统中的颗粒诱导电泳流，从而确定了我们最终将详细研究的初始流动参数体系。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一些未解决的问题的基本理解，这些问题直接影响到这些条形码阅读器的性能。这些问题包括加深我们对以下问题的理解：(A)棒状聚合物和棒状胶体颗粒悬浮液从稀释到半稀释的流变性，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相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作用；以及(D)非局部流动中棒状悬浮液的集体动力学，即平均流动尺度在棒材长度的量级上。事实上，尽管这些原理是复杂流体的微流体科学的固有原理，但这些非线性物理的许多组合将是第一次被研究。此外，通过与斯坦福大学科学教师暑期研究计划的合作，使用大规模计算设计微流控设备所产生的广泛教育影响将作为高中科学教师暑期实习的组成部分进行开发。这些实习将包括一两所TITLI学校的教师。一次实习将与研究的实验方面有关，另一次与计算设计方面有关。实习将允许教员与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.