这项工作得到了科学与台湾技术下的授权号101-2221-E-002-064-MY3部的支持。

1.制造微流体装置<ol><li>使用图形排版软件（ 如 AutoCAD的）画一个T-微通道的轮廓。为T形微通道中，两个进料通道90微米宽和2500微米长，合流通道为180μm宽和3000微米长。每个通道的一端连接到一个单独的圆的直径为1100微米。 </li><li>马克"清除"和"黑暗"的曝光和覆盖的区域，分别。对于负光致抗蚀剂（ 例如，SU-8）中，T-微通道的形状是"清除"和周围是"暗"。 </li><li>使用激光图案发生器具有442纳米的波长为2微米的最小特征尺寸，以T型微通道的图案转印到铬在玻璃上的光掩模。 </li><li>使用的光掩模，基板（ 例如，单面抛光的硅晶片）和永久环氧光致抗蚀剂（ 例如，SU-8），以通过一个标准的光刻工艺使一个模具。光致抗蚀剂层是55.2微米厚。在一般情况下，光致抗蚀剂厚度应比物镜17-19相关的深度薄。 </li><li>使用的模具和透明材料，例如聚二甲基硅氧烷（PDMS）来制造T型微通道20。 </li><li>用于流体连接的，使用2毫米的不锈钢管外径穿通孔对准PDMS上的圆形图案。 </li><li>治疗的PDMS和用氧等离子体于60W下30秒的玻璃载片的表面上。附加的PDMS到载玻片上。两种材料的氧化表面形成强大的粘合。将粘结的PDMS结构的热板上进行5分钟，在120℃。 </li><li>插入特氟隆管放置在冲压孔对流体连接。 </li></ol> 2.实验装置<ol><li>从Hoffma构建微型纹影系统n个调制对比度显微镜通过在冷凝器的前焦平面除去狭缝板和带刀口在5X物镜3的后焦平面替换调制器。相关的深度，这取决于目标17-19的数值孔径，应足以覆盖所述微流体器件的整个深度。刀口的表面通过阳极氧化铝变黑来降低其反射率。 </li><li>通过C型安装适配器安装高速摄像机显微镜的三目镜筒。有相机通过一个分束器朝向显微镜的光路中。通过以太网电缆将相机连接至一台台式电脑。相机设置的伽马校正为1，即它的灰度读数是正比于输入亮度。 </li><li>打开光源。为了避免过多的热量，使用LED（发光二极管）照明。 </li><li>使用图像处理软件（ 如&lt;/ em&gt;的在MATLAB函数imread），以获得所获取的图像的灰度级值。除去刀口，调整照明，光圈和曝光时间，以使图像的平均灰度读数比最大值低约10％。这表示背景强度为0％的截止和我们使用230的值对于8位图像。 </li><li>插入刀口完全阻断入射光。记录的图像的平均灰度读数。这表示本底密度为100％的截止和值约为15 8位图像。 </li><li>调整刀口的位置，使得所获取的图像的平均灰度读数在于为0％和100％的截止值的中间。现在截止的程度被设置为50％。 </li><li>准备两个透明流体与已知折射率21彼此作为组分完全混溶。为了评价屈光IND的依赖恩对混合物浓度，检查文献21或使用格莱斯顿，戴尔方程22。如果曲线是非线性的，在整个范围内，挑其它流体组分。然后，选择一个指定的组合物，低于该溶液的折射率与浓度线性变化。例如，使用稀的含水乙醇为0.05质量分数和水作为工作流体。 </li><li>把T型微通道上的试样台。安排T形微通道因此与平行于刀口合流通道（ 图1）。 </li><li>准备两个相同的注射器:注射器A被填充有工作流体在作为参考流体（水），和注射器B被填充有其它工作流体（稀乙醇水溶液）。注射器的大小取决于所需的流量Q和注射泵的规格:Q =πD2 V / 4，其中d是SYR的内径英格和V是柱塞的速度。流量脉动通常可以通过选择一个小的注射器，以增加V 23来防止。 </li><li>收集从T-微通道中的烧杯中的出口流体。确保特氟隆出口管固定到烧杯的壁和其端部是在液面以下，在烧杯中，以避免这将是由液滴片断离引起的振动。 </li></ol> 3.校准<ol><li>获得两种流体混合和参考图像的图像。 <ol><li>在给定的雷诺数 Re，设置注射泵的流速，Q。Q选自Q =μ（W + D）的重 /4ρ，其中，μ和ρ是粘度和工作流体的密度，和W计算和 D是T型微通道的合流通道的宽度和深度，分别。 </li><li>装入一张泵用注射器A和其他泵SYB. RINGE连接T-微通道的两个入口通过聚四氟乙烯管注射器和注射器湾启动注射泵输​​出工作流体到T-微通道以相同体积流量。 </li><li>等到源源不断建立。稳流状态是由一个固定的图案纹影的出现定义。 </li><li>使用相机控制软件来记录流体混合的20个帧在30帧每秒的帧率。 </li><li>停止其装入注射器B中的泵只有通过一个入口泵送参考流体（水）到T-微通道以恒定速率合流通道。 </li><li>等到稳流状态达到无条纹图案是观察。 </li><li>使用相机控制软件作为参考图像，当没有光学不均匀性存在于T形微通道。记录20个帧在30帧每秒的帧率。 </li><li> 3.1.1重复3.1.7到在不同的雷诺数: 重新 = 1，5，10，20和50，以便没有复杂流动结构出现在T-微通道24的合流区域。 </li></ol></li><li>使用图像处理软件由参考图像I 0（I，J）25，其中i和j是该像素指数来划分所获取的图像I（i，j）的 。 </li><li>采用CFD（计算流体动力学）封装，以模拟在T-微通道中的指定的流体的混合。 <ol><li>构造为T型微通道的几何形状的三维模型。离散流域划分为结构化网格。提高精度，采用更细的网眼的合流和T-微通道的中心区域。 </li><li>分配的流体的物理性质，并建立边界条件到流域。在求解过程中，确定从在最后获得的浓度的浓度依赖性的扩散系数迭代26，以更新的局部浓度。 </li><li>通过执行电网研究27检查的计算结果的敏感性。 </li></ol></li><li>对于每一个节点（X I，Y I）在xy -平面，应用CFD后处理工具，采取跨航道水深浓度场的平均值由梯形规则:W（X I，Y j）条 = {ΣK表 [ 瓦特（X I，YĴ和z k）的 + W（X I，YĴ和zķ1）]∙（Zķ1 - z k）的 / 2} / D 28，其中D是通道深度。使用中心差分格式来计算deriva集中略去相对于交叉流方向:（∂W /∂Y）I，J = [W（X I，YĴ+1） - W（X I，YĴ-1）] /（YĴ +1 - ÿĴ-1）。 </li><li>对于正的和负的梯度，提取灰度值I / I 0（在3.2中获得）和质量分数的梯度∂瓦特/∂ý在指定的地点，例如y = 0 的 （中心线（在3.4获得），流向的比率方向）或各种给定的x（横流方向）。 </li><li>画出的结果，并确定我之间的关系/ I 0和∂W /∂Y，I / I 0 </sub&gt;为C 1〜∂重量/∂Y + C 2（C 1和C 2是常数），用线性回归。 </li></ol> 4.定量<ol><li>重复步骤3.1至3.2中的目标微流体装置中混合。目标微流体装置的深度应该是相同或接近的T-微通道。如果不稳定现象有望，获取视频剪辑（图像序列）的步骤，而不是3.1.4。帧速率应该足够高，以解决瞬态流动明确的动态，而曝光时间应该是相同的，以在2.4，2.5，3.1.4和3.1.7中使用的值。 </li><li>使用在步骤3.6中得到的关系为灰度值的比率转换为质量分数的目标微流体装置的梯度。 </li></ol>

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Y. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Microscale+schlieren+visualization+of+near-bubble+mass+transport+during+boiling+of+2-propanol/water+mixtures+in+a+square+capillary.">Microscale schlieren visualization of near-bubble mass transport during boiling of 2-propanol/water mixtures in a square capillary.</a> Experiments in Fluids. 55 (7), 1778 (2014).</li><li>Panigrahi, P. K., Muralidhar, K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> Schlieren and Shadowgraph Methods in Heat and Mass Transfer. , (2012).</li></ol>

作者什么都没有透露。

用于在微流体装置的流体的混合，所述微尺度纹影技术能够通过在光强度量化变化来测量浓度梯度的大小。因为这种技术的原理依赖于探测光的传播的轮换时，工作流体和微流体装置必须是透明的入射光。此外，该协议要求的溶液，其组成，以使工作流体的初步评估是必不可少的折射率之间的线性关系。除了 ​​乙醇水溶液在此证明，微型纹影技术成功地应用于测量盐度梯度29和solutocapillary对流30。为了精确的测量，孔径范围，照明水平，曝光时间，物镜和微通道深度的校准过程中使用的必须是完全相同的定量过程中使用。此外，上述物镜的相关性的深度必须足够大以覆盖所述微流体器件的整个深度。 在T-微混合的校准过程是在微尺度纹影技术的准确定量最关键的一步。为了成功实施该方法，用户需要对齐管连接正确，利用小注射器或气动流体输送，以避免流动振荡23，使用LED光源，减少多余的热量，进行校准程序在低雷诺数24，并 ​​放置在微流体装置中的焦点，以消除高阶光学效应31。最低可测量梯度（亮模式，∂重量/∂是 &lt;0）被连接到摄像机的动态范围，而最高可测量梯度（暗图案，∂瓦特/∂ÿ&gt; 0）时达到刀口完全阻断偏转光。以检测多种浓度梯度的，高的ISO值是有利的，只要曝光不足或曝光过度，不会发生。方法的检出限，低于该微纹影系统无法识别，依赖于最小强度的变化，相机能够解决。最小强度变化是由噪声的程度和色调层次的水平的限制。因此，高灵敏度照相机以极大的像素深度期望为低信号的应用程序。 微尺度纹影技术的意义在于两个折叠;一方面，它使非定常满场测量实时地与简单的光学构造。另一方面，它是无创使得没有外来物质被引入到扰乱流场。因为微纹影技术产生三维不均匀的二维投影在一个微法luidic设备，复杂的混合现象仍然隐晦通过现有的方法可以清楚地看到。这种技术的未来应用包括在电化学过程量化浓度梯度或确定营养梯度来研究微生物的趋化性在微流环境。

流体混合]是在许多工业过程和生物系统中发现的一个重要问题。随着微流体的出现，在微型混合带来了太多的关注，因为大众运输机制之间的扩散统治的挑战。由于设计一个有效所需的定量微混合器验证，几种测量方法被开发5-7。然而，该三维结构中，在有效的微混合器5通常发现，要求的共同测量技术不能提供浓度场的更精确的表示。由于观看角度8或反应动力学6的限制，上述方法可产生不正确地占混合物的均匀误导的结果。 对于光学透明的液体光学透明的微观混合，微型纹影技术3,9-14 </SUP&gt;提供了一个有吸引力的替代分析混合不均匀。在过去，微型纹影技术已主要用于可视可压缩流动9-13，15或相位梯度16。从两个简单的光学布局和高灵敏度的微型纹影技术的好处，能够实现特定的流动特性，使得光学干扰而非常适合于在评估混合使用不仅非侵入性的调查。在本文中，我们通过将刀口在显微镜的物镜的后焦平面构建微尺度纹影系统，描述的校准步骤，以实现定量分析，并在微流体振荡器4报告验证测量。为了实现测量时，工作流体被适当地选择，使得混合的流体的折射率与该组合物线性地变化，并且目标的微流体装置的厚度是相同的上ê用于校准。此外物种浓度，微量纹影技术可以扩展到测量其它标量是线性相关的折射率，如温度或盐度的梯度。

<table><tbody><tr><td>Permanent Epoxy Negative Photoresist</td><td>MicroChem</td><td>SU-8 2150</td><td></td></tr><tr><td>single side polished silicon wafer</td><td>Light Technology</td><td>S4W1PP5SABUP1</td><td>p-type, &lt;br /&gt;diameter: 100&amp;plusmn;0.5 mm, thickness: 525&amp;plusmn;25 mm, orientation: (1 0 0)</td></tr><tr><td>syringe pump</td><td>kdScientific</td><td>kds210</td><td></td></tr><tr><td>syringe, &lt;em&gt;i.e.&lt;/em&gt; 10 ml</td><td>Terumo</td><td>SS-10L2138</td><td></td></tr><tr><td>Hoffman modulation contrast microscope</td><td>Leica Microsystems</td><td>DM IL LED</td><td></td></tr><tr><td>5X objective lens</td><td>Leica Microsystems</td><td>N PLAN</td><td>NA = 0.12</td></tr><tr><td>knife-edge</td><td></td><td></td><td>custom made part</td></tr><tr><td>camera, &lt;em&gt;i.e.&lt;/em&gt; high speed</td><td>Integrated Design Tools</td><td>NX7-S1</td><td></td></tr><tr><td>C-mount adapter HC 0.63x</td><td>Leica Microsystems</td><td>541537</td><td></td></tr><tr><td>camera operating software</td><td>Integrated Design Tools</td><td>MotionPro X Studio 2.02.01</td><td></td></tr><tr><td>polydimethylsiloxane (PDMS)</td><td>Dow Corning</td><td>Sylgard-184 silicone elastomer kit</td><td></td></tr><tr><td>Teflon tubing, &lt;em&gt;i.e.&lt;/em&gt; O.D. x I.D. 1/16 in. x 0.031 in.</td><td>Supelco</td><td>58700-U</td><td></td></tr><tr><td>micro glass slide</td><td>Matsunami Glass</td><td>S2215</td><td></td></tr><tr><td>hot plate</td><td>Yeong-Shin</td><td>HP-303DN</td><td></td></tr><tr><td>distilled water, &lt;em&gt;i.e.&lt;/em&gt; HPLC grade</td><td>Alps Chemicals</td><td></td><td></td></tr><tr><td>ethyl alcohol, &lt;em&gt;i.e.&lt;/em&gt; reagent grade</td><td>Nihon Shiyaku Reagent</td><td>EA448652</td><td></td></tr><tr><td>image processing software</td><td>Mathworks</td><td>MATLAB R2009a</td><td></td></tr><tr><td>computational fluid dynamics package</td><td>ESI group</td><td>CFD-ACE+ 2008</td><td></td></tr></tbody></table>

analyzing mixing inhomogeneity in a microfluidic device by microscale schlieren technique

在本文中，我们介绍了利用微型纹影技术来衡量一个微流体装置的混合不均匀。微尺度纹影系统从霍夫曼调节对比度显微镜，它提供了容易获得的物镜的后焦平面构造，通过除去狭缝板和带刀口替换调制器。微尺度纹影技术的工作原理依赖于检测由折射率1-3的变化的光偏转。偏转光要么逃逸或被刀口受阻，产生明亮或暗带，分别为。如果该混合物的折射率随其组成而线性变化，在光强度在图像平面中的局部变化是正比于浓度梯度垂直于光轴。微纹影图像为按三维不均匀性产生的干扰光的二维投影。 为了完成定量分析，我们描述了在T形微通道混合两种流体的校准步骤。我们进行了数值模拟，得到在T-微通道，与相应的微纹影图像相关密切的浓度梯度。通过比较，微纹影图像的灰度读数和在微流体设备呈现的浓度梯度之间的关系被建立。使用这种关系，我们能够分析来自准微纹影图像的混合不均匀性，并证明微尺度纹影技术的能力与测量微流体振荡器4。用于光学透明流体，微尺度纹影技术是一个有吸引力的诊断工具，以提供其保留的混合过程的三维特征的瞬时满场信息。

灰度比I / I 0 的下不同雷诺数为正的和负的质量分数的梯度示出（ 图2）具有对称带出现在T型微通道的中间。在低雷诺数，纹影带的尾部被扩展并模糊由于横跨混合界面的分散。雷诺数增加时，扩散长度缩短导致更窄的频带。在不同的下游位置，强度变化ΔI的变化量/ I 0沿着横流方向定量描绘（ 图3）。从校准过程的结果被表示（ 图4A 和4B）。 我之间的关系/ I 0和∂W /∂y是线性雷诺NUM和独立BER。从回归分析中，I / I 0 = -110∂W /∂Y + 1.03∂W /∂Y&gt; 0，I / I 0 = -160∂W /∂Y + 0.83∂W /∂是 &lt;0 ，∂瓦特/∂y是微米-1。相对不确定度为±3.8％和±3.2％在图4A和4B分别。达到检测极限，其中数据点水平了。值得注意的是，在正，负梯度的斜坡的偏差的情况并不少见3。使用这些方程，质量分数梯度的随时间的变化在微流体振荡器4看出（ 图5）。混合接口被偏转的谐振区域和流动不稳定性Çommences。该视频图清楚地揭示了在微流体振荡器的流动的振荡性质，并演示微尺度纹影技术的能力来捕捉时间分辨全视野的浓度梯度在一个微流体装置。 <img alt="图1" src="/files/ftp_upload/52915/52915fig1.jpg" /> 图1示意图的光学设置的。刀口的取向产生暗带与折射率的正梯度。光偏转朝向增加折射率的方向。由于物镜反转图像，阻断- Ÿ区域屏蔽了扭曲的光，并产生一条暗带。 <img alt="图2" src="/files/ftp_upload/52915/52915fig2.jpg" /> 图2.比率灰度读数的用于在T-微混合理解过程<html> - [R不同的流量配置，正和，分别负梯度导致黑暗和明亮带。随着雷诺数的增加，带变得更集中。 <img alt="图3" src="/files/ftp_upload/52915/52915fig3.jpg" /> 图3.沿着横流方向上的强度变化为正的和负的梯度的变化。 再 = 1， 再 = 5。 <img alt="图4" src="/files/ftp_upload/52915/52915fig4.jpg" /> 图的质量分数的梯度和灰度比之间4.关系。对于正的和负的梯度，灰度比与质量分数梯度线性变化。 915 / 52915fig5.jpg"/&gt; <a href="https://www-jove-com.bdigitaluss.remotexs.co/files/ftp_upload/52915/52915fig5.mp4" target="_blank">图5（视频图）。</a>在微流体振荡器以雷诺数为 250通过流量振荡的混合特性演进质量分数梯度的成功由微型纹影技术捕获。

Watch this Scientific Journal Video about Analyzing Mixing Inhomogeneity in a Microfluidic Device by Microscale Schlieren Technique at JoVE.com

Analyzing Mixing Inhomogeneity in a Microfluidic Device by Microscale Schlieren Technique

Herein, we describe a procedure that employs microscale schlieren technique to measure mixing inhomogeneity in a microfluidic device. Through calibration, distribution of concentration gradient can be derived from the micro-schlieren image.

由微型条纹分析技术在微流控设备混合不均匀

In this paper, we introduce the use of microscale schlieren technique to measure mixing inhomogeneity in a microfluidic device. The microscale schlieren ...

analyzing-mixing-inhomogeneity-microfluidic-device-microscale

Universidad San Sebastian

Research

JoVE Journal

Bioengineering

8.9K 次观看.  National Taiwan University. Herein, we describe a procedure that employs microscale schlieren technique to measure mixing inhomogeneity in a microfluidic device. Through calibration, distribution of concentration gradient can be derived from the micro-schlieren image. 

视频: Analyzing Mixing Inhomogeneity in a Microfluidic Device by Microscale Schlieren Technique

Fabrication, Operation and Flow Visualization in Surface-acoustic-wave-driven Acoustic-counterflow Microfluidics

Surface acoustic waves (SAWs) can be used to drive liquids in portable microfluidic chips via the acoustic counterflow phenomenon. In this video we present the fabrication protocol for a multilayered SAW acoustic counterflow device. The device is fabricated starting from a lithium niobate (LN) substrate onto which two interdigital transducers (IDTs) and appropriate markers are patterned. A polydimethylsiloxane (PDMS) channel cast on an SU8 master mold is finally bonded on the patterned substrate. Following the fabrication procedure, we show the techniques that allow the characterization and operation of the acoustic counterflow device in order to pump fluids through the PDMS channel grid. We finally present the procedure to visualize liquid flow in the channels. The protocol is used to show on-chip fluid pumping under different flow regimes such as laminar flow and more complicated dynamics characterized by vortices and particle accumulation domains.

In this video we first describe fabrication and operation procedures of a surface acoustic wave (SAW) acoustic counterflow device. We then demonstrate an experimental setup that allows for both qualitative flow visualization and quantitative analysis of complex flows within the SAW pumping device.

制造，操作和表面声波驱动声逆流微流体流动可视化

Surface acoustic waves (SAWs) can be used to drive liquids in portable microfluidic chips via the acoustic counterflow phenomenon. In this video we ...

科研

工程学

Confocal Imaging of Confined Quiescent and Flowing Colloid-polymer Mixtures

The behavior of confined colloidal suspensions with attractive interparticle interactions is critical to the rational design of materials for directed assembly1-3, drug delivery4, improved hydrocarbon recovery5-7, and flowable electrodes for energy storage8. Suspensions containing fluorescent colloids and non-adsorbing polymers are appealing model systems, as the ratio of the polymer radius of gyration to the particle radius&#160;and concentration of polymer control the range and strength of the interparticle attraction, respectively. By tuning the polymer properties and the volume fraction of the colloids, colloid fluids, fluids of clusters, gels, crystals, and glasses can be obtained9.&#160;Confocal microscopy, a variant of fluorescence microscopy, allows an optically transparent and fluorescent sample to be imaged with high spatial and temporal resolution in three dimensions. In this technique, a small pinhole or slit blocks the emitted fluorescent light from regions of the sample that are outside the focal volume of the microscope optical system. As a result, only a thin section of the sample in the focal plane is imaged. This technique is particularly well suited to probe the structure and dynamics in dense colloidal suspensions at the single-particle scale: the particles are large enough to be resolved using visible light and diffuse slowly enough to be captured at typical scan speeds of commercial confocal systems10. Improvements in scan speeds and analysis algorithms have also enabled quantitative confocal imaging of flowing suspensions11-16,37.&#160;In this paper, we demonstrate confocal microscopy experiments to probe the confined phase behavior and flow properties of colloid-polymer mixtures. We first prepare colloid-polymer mixtures that are density- and refractive-index matched. Next, we report a standard protocol for imaging quiescent dense colloid-polymer mixtures under varying confinement in thin wedge-shaped cells. Finally, we demonstrate a protocol for imaging colloid-polymer mixtures during microchannel flow.

Confocal microscopy is used to image quiescent and flowing colloid-polymer mixtures, which are studied as model systems for attractive suspensions. Image analysis algorithms are used to calculate structural and dynamic metrics for the colloidal particles that measure changes due to geometric confinement.

密闭静态和流动胶体聚合物混合物的共聚焦成像

The behavior of confined colloidal suspensions with attractive interparticle interactions is critical to the rational design of materials for directed ...

化学

Microfluidic Chips Controlled with Elastomeric Microvalve Arrays

Miniaturized microfluidic systems provide simple and effective solutions for low-cost point-of-care diagnostics and high-throughput biomedical assays. Robust flow control and precise fluidic volumes are two critical requirements for these applications. We have developed microfluidic chips featuring elastomeric polydimethylsiloxane (PDMS) microvalve arrays that: 1) need no extra energy source to close the fluidic path, hence the loaded device is highly portable; and 2) allow for microfabricating deep (up to 1 mm) channels with vertical sidewalls and resulting in very precise features.

The PDMS microvalves-based devices consist of three layers: a fluidic layer containing fluidic paths and microchambers of various sizes, a control layer containing the microchannels necessary to actuate the fluidic path with microvalves, and a middle thin PDMS membrane that is bound to the control layer. Fluidic layer and control layers are made by replica molding of PDMS from SU-8 photoresist masters, and the thin PDMS membrane is made by spinning PDMS at specified heights. The control layer is bonded to the thin PDMS membrane after oxygen activation of both, and then assembled with the fluidic layer. The microvalves are closed at rest and can be opened by applying negative pressure (e.g., house vacuum). Microvalve closure and opening are automated via solenoid valves controlled by computer software.

Here, we demonstrate two microvalve-based microfluidic chips for two different applications. The first chip allows for storing and mixing precise sub-nanoliter volumes of aqueous solutions at various mixing ratios. The second chip allows for computer-controlled perfusion of microfluidic cell cultures.

The devices are easy to fabricate and simple to control. Due to the biocompatibility of PDMS, these microchips could have broad applications in miniaturized diagnostic assays as well as basic cell biology studies.

We demonstrate protocols for manufacturing and automating elastomeric polydimethylsiloxane (PDMS)-based microvalve arrays that need no extra energy to close and feature photolithographically defined precise volumes. A parallel subnanoliter-volume mixer and an integrated microfluidic perfusion system are presented.

微流控芯片控制，弹性微阀阵列

Miniaturized microfluidic systems provide simple and effective solutions for low-cost point-of-care diagnostics and high-throughput biomedical assays.  ...

生物学

Microfluidic Mixers for Studying Protein Folding

The process by which a protein folds into its native conformation is highly relevant to biology and human health yet still poorly understood. One reason for this is that folding takes place over a wide range of timescales, from nanoseconds to seconds or longer, depending on the protein1. Conventional stopped-flow mixers have allowed measurement of folding kinetics starting at about 1 ms. We have recently developed a microfluidic mixer that dilutes denaturant ~100-fold in ~8 &mu;s2. Unlike a stopped-flow mixer, this mixer operates in the laminar flow regime in which turbulence does not occur. The absence of turbulence allows precise numeric simulation of all flows within the mixer with excellent agreement to experiment3-4.
Laminar flow is achieved for Reynolds numbers Re &le;100. For aqueous solutions, this requires micron scale geometries. We use a hard substrate, such as silicon or fused silica, to make channels 5-10 &mu;m wide and 10 &mu;m deep (See Figure 1). The smallest dimensions, at the entrance to the mixing region, are on the order of 1 &mu;m in size. The chip is sealed with a thin glass or fused silica coverslip for optical access. Typical total linear flow rates are ~1 m/s, yielding Re~10, but the protein consumption is only ~0.5 nL/s or 1.8 &mu;L/hr. Protein concentration depends on the detection method: For tryptophan fluorescence the typical concentration is 100 &mu;M (for 1 Trp/protein) and for FRET the typical concentration is ~100 nM.
The folding process is initiated by rapid dilution of denaturant from 6 M to 0.06 M guanidine hydrochloride. The protein in high denaturant flows down a central channel and is met on either side at the mixing region by buffer without denaturant moving ~100 times faster (see Figure 2). This geometry causes rapid constriction of the protein flow into a narrow jet ~100 nm wide. Diffusion of the light denaturant molecules is very rapid, while diffusion of the heavy protein molecules is much slower, diffusing less than 1 &mu;m in 1 ms. The difference in diffusion constant of the denaturant and the protein results in rapid dilution of the denaturant from the protein stream, reducing the effective concentration of the denaturant around the protein. The protein jet flows at a constant rate down the observation channel and fluorescence of the protein during folding can be observed using a scanning confocal microscope5.

In this work we explain the fabrication and use of a microfluidic mixer capable of mixing two solutions in ~8 &mu;s. We also demonstrate the use of these mixers with spectroscopic detection using UV fluorescence and fluorescence resonance energy transfer (FRET).

研究蛋白质折叠的微流体混合器

The process by which a protein folds into its native conformation is highly relevant to biology and human health yet still poorly understood. One reason ...

生物工程

A Microfluidic Device for Studying Multiple Distinct Strains

The study of cell responses to environmental changes poses many experimental challenges: cells need to be imaged under changing conditions, often in a comparative manner. Multiwell plates are routinely used to compare many different strains or cell lines, but allow limited control over the environment dynamics. Microfluidic devices, on the other hand, allow exquisite dynamic control over the surrounding conditions, but it is challenging to image and distinguish more than a few strains in them. Here we describe a method to easily and rapidly manufacture a microfluidic device capable of applying dynamically changing conditions to multiple distinct yeast strains in one channel. The device is designed and manufactured by simple means without the need for soft lithography. It is composed of a Y-shaped flow channel attached to a second layer harboring microwells. The strains are placed in separate microwells, and imaged under the exact same dynamic conditions. We demonstrate the use of the device for measuring protein localization responses to pulses of nutrient changes in different yeast strains.

We present a simple method to produce microfluidic devices capable of applying similar dynamic conditions to multiple distinct strains, without the need for a clean room or soft lithography.

一种微流体装置研究多个不同的菌株

The study of cell responses to environmental changes poses many experimental challenges: cells need to be imaged under changing conditions, often in a ...

Combining Microfluidics and Microrheology to Determine Rheological Properties of Soft Matter during Repeated Phase Transitions

The microstructure of soft matter directly impacts macroscopic rheological properties and can be changed by factors including colloidal rearrangement during previous phase changes and applied shear. To determine the extent of these changes, we have developed a microfluidic device that enables repeated phase transitions induced by exchange of the surrounding fluid and microrheological characterization while limiting shear on the sample. This technique is &#181;2rheology, the combination of microfluidics and microrheology. The microfluidic device is a two-layer design with symmetric inlet streams entering a sample chamber that traps the gel sample in place during fluid exchange. Suction can be applied far away from the sample chamber to pull fluids into the sample chamber. Material rheological properties are characterized using multiple particle tracking microrheology (MPT). In MPT, fluorescent probe particles are embedded into the material and the Brownian motion of the probes is recorded using video microscopy. The movement of the particles is tracked and the mean-squared displacement (MSD) is calculated. The MSD is related to macroscopic rheological properties, using the Generalized Stokes-Einstein Relation. The phase of the material is identified by comparison to the critical relaxation exponent, determined using time-cure superposition. Measurements of a fibrous colloidal gel illustrate the utility of the technique. This gel has a delicate structure that can be irreversibly changed when shear is applied. &#181;2rheology data shows that the material repeatedly equilibrates to the same rheological properties after each phase transition, indicating that phase transitions do not play a role in microstructural changes. To determine the role of shear, samples can be sheared prior to injection into our microfluidic device. &#181;2rheology is a widely applicable technique for the characterization of soft matter enabling the determination of rheological properties of delicate microstructures in a single sample during phase transitions in response to repeated changes in the surrounding environmental conditions.

We demonstrate the fabrication and use of a microfluidic device that enables multiple particle tracking microrheology measurements to study the rheological effects of repeated phase transitions on soft matter.

微流体和 Microrheology 相结合测定软物质在重复相变过程中的流变特性

The microstructure of soft matter directly impacts macroscopic rheological properties and can be changed by factors including colloidal rearrangement ...

One-Step Approach to Fabricating Polydimethylsiloxane Microfluidic Channels of Different Geometric Sections by Sequential Wet Etching Processes

Polydimethylsiloxane (PDMS) materials are substantially exploited to fabricate microfluidic devices by using soft lithography replica molding techniques. Customized channel layout designs are necessary for specific functions and integrated performance of microfluidic devices in numerous biomedical and chemical applications (e.g., cell culture, biosensing, chemical synthesis, and liquid handling). Owing to the nature of molding approaches using silicon wafers with photoresist layers patterned by photolithography as master molds, the microfluidic channels commonly have regular cross sections of rectangular shapes with identical heights. Typically, channels with multiple heights or different geometric sections are designed to possess particular functions and to perform in various microfluidic applications (e.g., hydrophoresis is used for sorting particles and in continuous flows for separating blood cells6,7,8,9). Therefore, a great deal of effort has been made in constructing channels with various sections through multiple-step approaches like photolithography using several photoresist layers and assembly of different PDMS thin sheets. Nevertheless, such multiple-step approaches usually involve tedious procedures and extensive instrumentation. Furthermore, the fabricated devices may not perform consistently and the resulted experimental data may be unpredictable. Here, a one-step approach is developed for the straightforward fabrication of microfluidic channels with different geometric cross sections through PDMS sequential wet etching processes, that introduces etchant into channels of planned single-layer layouts embedded in PDMS materials. Compared to the existing methods for manufacturing PDMS microfluidic channels with different geometries, the developed one-step approach can significantly simplify the process to fabricate channels with non-rectangular sections or various heights. Consequently, the technique is a way of constructing complex microfluidic channels, which provides a fabrication solution for the advancement of innovative microfluidic systems.

Several methods are available for the fabrication of channels of non-rectangular sections embedded in polydimethylsiloxane microfluidic devices. Most of them involve multistep manufacturing and extensive alignment. In this paper, a one-step approach is reported for fabricating microfluidic channels of different geometric cross sections by polydimethylsiloxane sequential wet etching.

连续湿法刻蚀法制作不同几何断面烷微流控通道的一步法

Polydimethylsiloxane (PDMS) materials are substantially exploited to fabricate microfluidic devices by using soft lithography replica molding techniques. ...

A Microfluidic Platform for Precision Small-volume Sample Processing and Its Use to Size Separate Biological Particles with an Acoustic Microdevice

A major advantage of microfluidic devices is the ability to manipulate small sample volumes, thus reducing reagent waste and preserving precious sample. However, to achieve robust sample manipulation it is necessary to address device integration with the macroscale environment. To realize repeatable, sensitive particle separation with microfluidic devices, this protocol presents a complete automated and integrated microfluidic platform that enables precise processing of 0.15&#8211;1.5 ml samples using microfluidic devices. Important aspects of this system include modular device layout and robust fixtures resulting in reliable and flexible world to chip connections, and fully-automated fluid handling which accomplishes closed-loop sample collection, system cleaning and priming steps to ensure repeatable operation. Different microfluidic devices can be used interchangeably with this architecture. Here we incorporate an acoustofluidic device, detail its characterization, performance optimization, and demonstrate its use for size-separation of biological samples. By using real-time feedback during separation experiments, sample collection is optimized to conserve and concentrate sample. Although requiring the integration of multiple pieces of equipment, advantages of this architecture include the ability to process unknown samples with no additional system optimization, ease of device replacement, and precise, robust sample processing.

This protocol describes a system architecture for performing automated small volume (0.15&#8211;1.5 ml) particle separations using a microfluidic device, and discusses methods to optimize acoustofluidic device performance and operation.

微流体平台精密小批量来样加工及它的使用尺寸分离生物颗粒的声微型

A major advantage of microfluidic devices is the ability to manipulate small sample volumes, thus reducing reagent waste and preserving precious sample. ...

Window on a Microworld: Simple Microfluidic Systems for Studying Microbial Transport in Porous Media

Microbial growth and transport in porous media have important implications for the quality of groundwater and surface water, the recycling of nutrients in the environment, as well as directly for the transmission of pathogens to drinking water supplies. Natural porous media is composed of an intricate physical topology, varied surface chemistries, dynamic gradients of nutrients and electron acceptors, and a patchy distribution of microbes. These features vary substantially over a length scale of microns, making the results of macro-scale investigations of microbial transport difficult to interpret, and the validation of mechanistic models challenging. Here we demonstrate how simple microfluidic devices can be used to visualize microbial interactions with micro-structured habitats, to identify key processes influencing the observed phenomena, and to systematically validate predictive models. Simple, easy-to-use flow cells were constructed out of the transparent, biocompatible and oxygen-permeable material poly(dimethyl siloxane). Standard methods of photolithography were used to make micro-structured masters, and replica molding was used to cast micro-structured flow cells from the masters. The physical design of the flow cell chamber is adaptable to the experimental requirements: microchannels can vary from simple linear connections to complex topologies with feature sizes as small as 2 &mu;m. Our modular EcoChip flow cell array features dozens of identical chambers and flow control by a gravity-driven flow module. We demonstrate that through use of EcoChip devices, physical structures and pressure heads can be held constant or varied systematically while the influence of surface chemistry, fluid properties, or the characteristics of the microbial population is investigated. Through transport experiments using a non-pathogenic, green fluorescent protein-expressing Vibrio bacterial strain, we illustrate the importance of habitat structure, flow conditions, and inoculums size on fundamental transport phenomena, and with real-time particle-scale observations, demonstrate that microfluidics offer a compelling view of a hidden world.

Microfluidic devices can be used to visualize complex natural processes in real time and at the appropriate physical scales. We have developed a simple microfluidic device that mimics key features of natural porous media for studying growth and transport of bacteria in the subsurface.

窗口上的一个微观世界：简单的微流体系统为研究多孔介质中的微生物运输

Microbial growth and transport in porous media have important implications for the quality of groundwater and surface water, the recycling of nutrients in ...

Polydimethylsiloxane-polycarbonate Microfluidic Devices for Cell Migration Studies Under Perpendicular Chemical and Oxygen Gradients

This paper reports a microfluidic device made of polydimethylsiloxane (PDMS) with an embedded polycarbonate (PC) thin film to study cell migration under combinations of chemical and oxygen gradients. Both chemical and oxygen gradients can greatly affect cell migration in vivo; however, due to technical limitations, very little research has been performed to investigate their effects in vitro. The device developed in this research takes advantage of a series of serpentine-shaped channels to generate the desired chemical gradients and exploits a spatially confined chemical reaction method for oxygen gradient generation. The directions of the chemical and oxygen gradients are perpendicular to each other to enable straightforward migration result interpretation. In order to efficiently generate the oxygen gradients with minimal chemical consumption, the embedded PC thin film is utilized as a gas diffusion barrier. The developed microfluidic device can be actuated by syringe pumps and placed into a conventional cell incubator during cell migration experiments to allow for setup simplification and optimized cell culture conditions. In cell experiments, we used the device to study migrations of adenocarcinomic human alveolar basal epithelial cells, A549, under combinations of chemokine (stromal cell-derived factor, SDF-1&#945;) and oxygen gradients. The experimental results show that the device can stably generate perpendicular chemokine and oxygen gradients and is compatible with cells. The migration study results indicate that oxygen gradients may play an essential role in guiding cell migration, and cellular behavior under combinations of gradients cannot be predicted from those under single gradients. The device provides a powerful and practical tool for researchers to study interactions between chemical and oxygen gradients in cell culture, which can promote better cell migration studies in more in vivo-like microenvironments.

The control of chemical and oxygen gradients is essential for cell cultures. This paper reports a polydimethylsiloxane-polycarbonate (PDMS-PC) microfluidic device capable of reliably generating combinations of chemical and oxygen gradients for cell migration studies, which can be practically utilized in biological labs without sophisticated instrumentation.

细胞迁移研究在垂直化学和氧梯度聚二甲基硅氧烷的聚碳酸酯微流体装置

This paper reports a microfluidic device made of polydimethylsiloxane (PDMS) with an embedded polycarbonate (PC) thin film to study cell migration under ...