我们感谢Lindback基金会的资金支持，而亚历山大西多连科医 ​​生和埃尔萨楚的富有成果的讨论和技术援助，以及罗伯特·史密斯教授的协助C.线虫检测。

注:在下面描述的步骤，实验室安全准则必须始终遵循。特别重要的是，当与高电压（&gt; 500 V）和处理化学品处理的安全性。 1，涂料的导电性基板与蜡烛的烟灰<ol><li>切割金属铜为矩形（75×43毫米，厚0.5毫米）。清洁浸没在铜蚀刻剂的铜基板为大约30秒，洗净用自来水进行约20秒，干燥的纸。 注意:如果使用方法1以下，更改尺寸为75×25毫米到适合的机器。 </li><li>扫铜衬底下的点燃石蜡蜡烛为30-45秒，以获得近似均匀的烟灰涂层（约40μm厚）。保持基板在约1cm焰内。请勿触摸脆弱的烟灰表面。 </li></ol> 2.保护与涂层的烟灰层注:烟灰层非常脆弱的，必须涂上保护层。两个简单的替代品（方法1和2所示）被认为在这里，但更可靠的协议，目前正在开发中。 <ol><li>方法1 <ol><li>加载样品放入金属蒸发或溅射系统。以下所述系统的操作过程，排空腔室，并且开始金受控沉积在烟灰层（150-200纳米）。让该装置冷却到室温。 </li><li>浸涂金属化基片在一个1-十二硫醇溶液（1％体积/体积，在95％乙醇中，ACS / USP级），用于化学罩内10分钟。然后，拿着该设备以一定的角度接近60°，轻轻地用只有几个乙醇滴洗表面。让设备干燥过夜。 </li></ol></li><li>方法2 <ol><li>在化学罩，涂敷煤烟基板后立即和同时衬底仍从蜡烛火焰温，沉积氟化液体的一些液滴上的一侧基板上，并倾斜衬底的角度接近90°。存放更多的水滴，让他们滚在整个油烟面。 注:当水滴落在一个点，烟灰会被冲离该区域。让的氟化液体扩散的液滴尽可能。 </li><li>烘烤的热板上（160℃，15分钟）的化学罩内的衬底。 </li><li>让基板放置过夜，在室温下在使用前。保存下去。 </li></ol></li></ol> 3.上电极的制作（从Abdelgawad 等人改编。12） <ol><li>通过绘制图形设计软件的电极。每个电极为2毫米长，0.3毫米宽，与电极之间的间隙为0.3mm。触头（卡入连接器中，见下文）之间的间隙为2.3毫米（ 图1）。 </li><li>修剪灵活的铜层压板（35微米厚）为国君格式（3.87×7.5cm的英寸）。使用其他尺寸的我f相的打印机兼容。加载层压到一台彩色打印机的手动进纸托盘。 </li><li>请务必使用"富黑"，或"登记黑"，在铜表打印时（见Abdelgawad 等 12查看详情），让黑色墨水的一层致密的铜基材上，蚀刻过程中保护印刷图案。让印刷基板完全干燥，过夜。 </li><li>内部的化学罩，预热（40℃），用50ml的铜蚀刻液的烧杯中。浸在烧杯中的印刷层压板，并轻轻摇动它在溶液中约10分钟。刻蚀时间取决于铜蚀刻剂溶液中。每隔几分钟，检查腐蚀，看看图案是否完好。 </li><li>小心地用水冲洗所述层压体，并用丙酮和乙醇中的化学引擎盖除去包衣。再次洗净，并轻轻擦干用纸巾层压板。 </li><li>小心地连接层压用电极为GLASS滑动件（75×25毫米，约1毫米厚），用双面胶带。避免气穴。 </li><li>附上氟烷的PFA薄膜，以使用磁带的电极。这用于防止电极与液滴的意外接触，从而损害顶电极由于短路。 </li></ol> 4.电子接口（电路如图2） <ol><li>焊接继电器和电容器C为通用电路板。 </li><li>装配了10继电器驱动器的其余部分在一个无焊面包板的电子电路。 </li><li>线各继电器驱动器的输入，以在所述控制板上的频道。 </li><li>小心管理单元的顶部电极到一个连接器（ 图3）。线的每个中继驱动器的输出到一个顶部电极，如该图所示。注意，有一对导线由继电器之间的接地连接器的接触，以最小化电噪声。 注:该连接器位于一个可调节的平台来控制T ON他的顶部和底部（碳黑涂覆）衬底之间的距离（0.1-0.5毫米）。 </li><li>使用程序来控制高电压（HV）的应用程序（约0.8秒），以4个电极在同一时间定时，移位1电极在运动（ 即方向，为0.8秒时，致动1234，然后2345，3456 等。，0.8秒的每个组，然后向后，使液滴移动在相反的方向上也同样）。 </li></ol> 5.滴可视化和处理<ol><li>以记录液滴的运动，可以使用可视化系统，它是由一个24X的 - 96X放大率组件结合了CCD照相机。视频记录器连接到使用S视频相机。 </li><li>吸管含C的 4微升滴线虫中的烟灰的基板的底部的介质。 </li><li>使顶端电极，以-0.3毫米的液滴之上。液滴应当接近中间，只是在第五电极的下方，以便于操作。 </li><li>把电子接口和高电压（500伏RMS）上，并调整顶部电极距离的液滴，直到它开始移动。不要让顶部电极接触到液滴。 </li><li>通过记录响应于电脉冲在设备上成功液滴转移的次数采集数据。一个成功的实验的特征在于，至少700微滴转移， 也就是说 ，每一个电脉冲后一个传送。 </li><li>连续地收集数据，直到微滴不响应再移动到5至10个脉冲。 注意:当表面开始退化，运动可以通过使顶电极更接近液滴被恢复。 </li></ol>

<ol>
	<li>Fair, R. B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Digital+microfluidics:+is+a+true+lab-on-a-chip+possible.">Digital microfluidics: is a true lab-on-a-chip possible.</a> Microfluid Nanofluid. 3 (3), 245-281 (2007).</li><li>Gupta, S., Alargova, R. G., Kilpatrick, P. K., Velev, O. D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=On-Chip+Dielectrophoretic+Coassembly+of+Live+Cells+and+Particles+into+Responsive+Biomaterials.">On-Chip Dielectrophoretic Coassembly of Live Cells and Particles into Responsive Biomaterials.</a> Langmuir. 26 (5), 3441-3452 (2009).</li><li>Shih, S. C., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Dried+blood+spot+analysis+by+digital+microfluidics+coupled+to+nanoelectrospray+ionization+mass+spectrometry.">Dried blood spot analysis by digital microfluidics coupled to nanoelectrospray ionization mass spectrometry.</a> Anal Chem. 84 (8), 3731-3738 (2012).</li><li>Gorbatsova, J., Borissova, M., Kaljurand, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Electrowetting-on-dielectric+actuation+of+droplets+with+capillary+electrophoretic+zones+for+off-line+mass+spectrometric+analysis.">Electrowetting-on-dielectric actuation of droplets with capillary electrophoretic zones for off-line mass spectrometric analysis.</a> J Chromatogr. 1234, 9-15 (2012).</li><li>Qin, J., Wheeler, A. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Maze+exploration+and+learning+in+C.+elegans.">Maze exploration and learning in C. elegans.</a> Lab Chip. 7 (2), 186-192 (2007).</li><li>Koc, Y., de Mello, A. J., McHale, G., Newton, M. I., Roach, P., Shirtcliffe, N. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Nano-scale+superhydrophobicity:+suppression+of+protein+adsorption+and+promotion+of+flow-induced+detachment.">Nano-scale superhydrophobicity: suppression of protein adsorption and promotion of flow-induced detachment.</a> Lab Chip. 8 (4), 582-586 (2008).</li><li>Perry, G., Thomy, V., Das, M. R., Coffinier, Y., Boukherroub, R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Inhibiting+protein+biofouling+using+graphene+oxide+in+droplet-based+microfluidic+microsystems.">Inhibiting protein biofouling using graphene oxide in droplet-based microfluidic microsystems.</a> Lab Chip. 12 (9), 1601-1604 (2012).</li><li>Kumari, N., Garimella, S. V. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Electrowetting-Induced+Dewetting+Transitions+on+Superhydrophobic+Surfaces.">Electrowetting-Induced Dewetting Transitions on Superhydrophobic Surfaces.</a> Langmuir. 27 (17), 10342-10346 (2011).</li><li>Freire, S. L. S., Tanner, B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Additive-Free+Digital+Microfluidics.">Additive-Free Digital Microfluidics.</a> Langmuir. 29 (28), 9024-9030 (2013).</li><li>Deng, X., Mammen, L., Butt, H. -. J., Vollmer, D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Candle+Soot+as+a+Template+for+a+Transparent+Robust+Superamphiphobic+Coating.">Candle Soot as a Template for a Transparent Robust Superamphiphobic Coating.</a> Science. 335, 67-70 (2011).</li><li>Kang, K. H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=How+Electrostatic+Fields+Change+Contact+Angle+in+Electrowetting.">How Electrostatic Fields Change Contact Angle in Electrowetting.</a> Langmuir. 18 (26), 10318-10322 (2002).</li><li>Abdelgawad, M., Watson, M. W. L., Young, E. W. K., Mudrik, J. M., Ungrin, M. D., Wheeler, A. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Soft+lithography:+masters+on+demand.">Soft lithography: masters on demand.</a> Lab Chip. 8 (8), 1379-1385 (2008).</li><li>Barbulovic-Nad, I., Yang, H., Park, P. S., Wheeler, A. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Digital+microfluidics+for+cell-based+assays.">Digital microfluidics for cell-based assays.</a> Lab Chip. 8 (4), 519-526 (2008).</li><li>Brenner, S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+genetics+of+Caenorhabditis+elegans.">The genetics of Caenorhabditis elegans.</a> Genetics. 77 (1), 71-94 (1974).</li></ol>

The authors declare that they have no competing financial interests.

该协议中最关键的步骤是保护的烟灰层的，可直接与在移动液滴的成功相关联。金属化的烟灰层（上述的方法1）允许接近制造成功的100％。然而，最大的操作时间约为10分钟;可能的话，微滴级分通过在所述金属层的孔润湿烟灰。涂烟灰层的氟化液是最简单和最快的替代，需要最少的资源，但只有40％-50％的基板制作工作（20分最高）的 - 和涂层不均匀。实际上，该烟灰层是非常脆弱的，而且在粘?...

该液体的工作装置的小型化是非常重要的用于"芯片实验室"的平台的开发。在这个方向上，在过去二十年已经见证了在微流体领域的显著进展，具有各种应用。1-5用流体在封闭的通道（通道微流）的传输对比，DMF操纵上的电极阵列的微滴。其中，最有吸引力的这种技术的优点是没有可动部件运送流体，并且运动是通过关闭电信号即刻停止。 然而，雾滴的运动依赖于液滴内容，当然对于通用的"芯片实验室"的平台不希望的特性。含有蛋白质和其它分析物的微滴粘到设备的表面，成为不可移动。可以说，这一直是主要的限制为拓宽DMF的应用范围; 6-8替代方案，以尽量减少不希望的表面结垢涉及添加额外的化学物质，以液滴或它的周围，这有可能影响液滴的内容。 先前，我们小组开发出一种装置，以允许细胞和蛋白质在DMF中的传输，而无需额外的添加剂（现场DW的设备）。9这是通过基于蜡烛烟灰，10与设备的几何形状有利于液滴滚动表面相结合来实现并导致在熔滴上的向上的力，从而进一步降低液滴的表面相互作用。在这种方法中，液滴运动并不与表面润湿相关联。11 下面描述的详细方法的目标是产生一个微流体装置能够输送含有蛋白质，细胞和整个生物体的液滴，而无需额外的添加剂。在现场-DW设备铺平了道路液滴工作的化学家很大程度上独立完全控制平台RY。 在这里，我们还存在模拟表明，尽管所需的设备的操作的高电压，在整个液滴上的电压降是所施加的电压的一小部分，这表明在液滴内部bioanalytes的影响可以忽略。事实上，与秀丽隐杆线虫 （ 线虫 ），用于各种生物学研究线虫的初步试验，表明蠕虫游泳原状作为施加电压。

<table><tbody><tr><td>Paraffin candle</td><td></td><td></td><td>Any paraffin candle</td></tr><tr><td>Sputtering system</td><td>Denton Vacuum, Moorestown, NJ</td><td></td><td>Sputter coater Desk V HP equipped with an Au target.&amp;nbsp;</td></tr><tr><td>1-dodecanethiol</td><td>Sigma-Aldrich</td><td>471364</td><td></td></tr><tr><td>Teflon</td><td>Dupont</td><td>AF-1600</td><td></td></tr><tr><td>Fluorinert FC-40</td><td>Sigma-Aldrich</td><td>F9755</td><td>Fluorinated liquid: Prepare Teflon-AF resin in Fluorinert FC-40, 1:100 (w/w), to create the hydrophobic coating.</td></tr><tr><td>Graphic design software -Adobe Illustrator</td><td>Adobe Systems</td><td></td><td>Other softwares might be used as well.</td></tr><tr><td>Copper laminate</td><td>Dupont</td><td>LF9110</td><td></td></tr><tr><td>Laser Printer</td><td>Xerox</td><td>Phaser 6360 or similar</td><td>Check for the compatibility with &quot;rich black&quot; or &quot;registration black&quot; (see text).</td></tr><tr><td>Copper Etchant</td><td>Transene</td><td>CE-100</td><td></td></tr><tr><td>Perfluoroalkoxy (PFA) film</td><td>McMaster-Carr</td><td>84955K22</td><td></td></tr><tr><td>Breadboard</td><td>Allied Electronics</td><td>70012450 or similar</td><td>Large enough to allow the assemble of 10 drivers.</td></tr><tr><td>Universal circuit board</td><td>Allied Electronics</td><td>70219535 or similar</td><td></td></tr><tr><td>Connector</td><td>Allied Electronics</td><td>5145154-8 or similar</td><td></td></tr><tr><td>Control board and control program (LabView software)</td><td>National Instruments</td><td>NI-6229 or similar</td><td></td></tr><tr><td>High-voltage amplifier</td><td>Trek</td><td>PZD700</td><td></td></tr><tr><td>Capacitors C and C1, 100 nF, 60 V</td><td>Allied&amp;nbsp;</td><td>8817183</td><td></td></tr><tr><td>Transistor T, NPN</td><td>Allied&amp;nbsp;</td><td>9350289</td><td></td></tr><tr><td>Diode D, 1N4007</td><td>Allied&amp;nbsp;</td><td>2660007</td><td></td></tr><tr><td>Relay&amp;nbsp;</td><td>Allied&amp;nbsp;</td><td>8862527</td><td></td></tr><tr><td>Visualization system</td><td>Edmund Optics</td><td>VZM 200i or similar</td><td>System magnification 24X &amp;#8211; 96X. It is combined with a Hitachi KP-D20B 1/2 in CCD Color Camera.</td></tr><tr><td>Recorder</td><td>Sony</td><td>GV-D1000 NTSC or similar</td><td>It is connected to the camera by an S-video cable.</td></tr><tr><td>Simulations</td><td>COMSOL Multiphysics</td><td>V. 4.4</td><td></td></tr></tbody></table>

taking advantage of reduced droplet-surface interaction to optimize transport of bioanalytes in digital microfluidics

Digital microfluidics (DMF), a technique for manipulation of droplets, is a promising alternative for the development of &#8220;lab-on-a-chip&#8221; platforms. Often, droplet motion relies on the wetting of a surface, directly associated with the application of an electric field; surface interactions, however, make motion dependent on droplet contents, limiting the breadth of applications of the technique.
Some alternatives have been presented to minimize this dependence. However, they rely on the addition of extra chemical species to the droplet or its surroundings, which could potentially interact with droplet moieties. Addressing this challenge, our group recently developed Field-DW devices to allow the transport of cells and proteins in DMF, without extra additives.
Here, the protocol for device fabrication and operation is provided, including the electronic interface for motion control. We also continue the studies with the devices, showing that multicellular, relatively large, model organisms can also be transported, arguably unaffected by the electric fields required for device operation.

以前，我们已经使用现场DW的设备，以允许蛋白质在DMF中的运动。特别是，用牛血清白蛋白（BSA）的液滴可以在一个浓度2000倍高于先前报道的其他作者（无添加剂）移动。这是由于在液滴和表面之间的相互作用减小; 图4显示了含有荧光标记的牛血清白蛋白的液滴（见弗莱雷等 9为在实验的详细信息）。在左侧的第一图为液滴坐在烟灰涂覆的表面;中间的一个，电场...

Watch this Scientific Journal Video about Taking Advantage of Reduced Droplet-surface Interaction to Optimize Transport of Bioanalytes in Digital Microfluidics at JoVE.com

Taking Advantage of Reduced Droplet-surface Interaction to Optimize Transport of Bioanalytes in Digital Microfluidics

The protocol for fabrication and operation of field dewetting devices (Field-DW) is described, as well as the preliminary studies of the effects of electric fields on droplet contents.

以减少液滴表面的相互作用来优化Bioanalytes运输中的数字微流体

Digital microfluidics (DMF), a technique for manipulation of droplets, is a promising alternative for the development of &#8220;lab-on-a-chip&#8221; ...

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7.8K 次观看.  University of the Sciences. The protocol for fabrication and operation of field dewetting devices (Field-DW) is described, as well as the preliminary studies of the effects of electric fields on droplet contents.

视频: Taking Advantage of Reduced Droplet-surface Interaction to Optimize Transport of Bioanalytes in Digital Microfluidics

Fluorescence detection methods for microfluidic droplet platforms

The development of microfluidic platforms for performing chemistry and biology has in large part been driven by a range of potential benefits that accompany system miniaturisation. Advantages include the ability to efficiently process nano- to femoto- liter volumes of sample, facile integration of functional components, an intrinsic predisposition towards large-scale multiplexing, enhanced analytical throughput, improved control and reduced instrumental footprints.1
In recent years much interest has focussed on the development of droplet-based (or segmented flow) microfluidic systems and their potential as platforms in high-throughput experimentation.2-4 Here water-in-oil emulsions are made to spontaneously form in microfluidic channels as a result of capillary instabilities between the two immiscible phases. Importantly, microdroplets of precisely defined volumes and compositions can be generated at frequencies of several kHz. Furthermore, by encapsulating reagents of interest within isolated compartments separated by a continuous immiscible phase, both sample cross-talk and dispersion (diffusion- and Taylor-based) can be eliminated, which leads to minimal cross-contamination and the ability to time analytical processes with great accuracy. Additionally, since there is no contact between the contents of the droplets and the channel walls (which are wetted by the continuous phase) absorption and loss of reagents on the channel walls is prevented.
Once droplets of this kind have been generated and processed, it is necessary to extract the required analytical information. In this respect the detection method of choice should be rapid, provide high-sensitivity and low limits of detection, be applicable to a range of molecular species, be non-destructive and be able to be integrated with microfluidic devices in a facile manner. To address this need we have developed a suite of experimental tools and protocols that enable the extraction of large amounts of photophysical information from small-volume environments, and are applicable to the analysis of a wide range of physical, chemical and biological parameters. Herein two examples of these methods are presented and applied to the detection of single cells and the mapping of mixing processes inside picoliter-volume droplets. We report the entire experimental process including microfluidic chip fabrication, the optical setup and the process of droplet generation and detection.

Droplet-based microfluidic platforms are promising candidates for high throughput experimentation since they are able to generate picoliter, self-compartmentalized vessels inexpensively at kHz rates. Through integration with fast, sensitive and high resolution fluorescence spectroscopic methods, the large amounts of information generated within these systems can be efficiently extracted, harnessed and utilized.

荧光检测方法，为微流体液滴平台

The development of microfluidic platforms for performing chemistry and biology has in large part been driven by a range of potential benefits that ...

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生物工程

A Microfluidic Chip for ICPMS Sample Introduction

This protocol discusses the fabrication and usage of a disposable low cost microfluidic chip as sample introduction system for inductively coupled plasma mass spectrometry (ICPMS). The chip produces monodisperse aqueous sample droplets in perfluorohexane (PFH). Size and frequency of the aqueous droplets can be varied in the range of 40 to 60 &#181;m and from 90 to 7,000&#160;Hz, respectively. The droplets are ejected from the chip with a second flow of PFH and remain intact during the ejection. A custom-built desolvation system removes the PFH and transports the droplets into the ICPMS. Here, very stable signals with a narrow intensity distribution can be measured, showing the monodispersity of the droplets. We show that the introduction system can be used to quantitatively determine iron in single bovine red blood cells. In the future, the capabilities of the introduction device can easily be extended by the integration of additional microfluidic modules.

We present a discrete droplet sample introduction system for inductively coupled plasma mass spectrometry (ICPMS). It is based on a cheap and disposable microfluidic chip that generates highly monodisperse droplets in a size range of 40&#8722;60 &#181;m at frequencies from 90 to 7,000 Hz.

微流体芯片ICPMS进样

This protocol discusses the fabrication and usage of a disposable low cost microfluidic chip as sample introduction system for inductively coupled plasma ...

A Pipette-Tip Based Method for Seeding Cells to Droplet Microfluidic Platforms

Amongst various microfluidic platform designs frequently used for cellular analysis, droplet-microfluidics provides a robust tool for isolating and analyzing cells at the single-cell level by eliminating the influence of external factors on the cellular microenvironment. Encapsulation of cells in droplets is dictated by the Poisson distribution as a function of the number of cells present in each droplet and the average number of cells per volume of droplet. Primary cells, especially immune cells, or clinical specimens can be scarce and loss-less encapsulation of cells remains challenging. In this paper, we present a new methodology that uses pipette-tips to load cells to droplet-based microfluidic devices without the significant loss of cells. With various cell types , we demonstrate efficient cell encapsulation in droplets that closely corresponds to the encapsulation efficiency predicted by the Poisson distribution. Our method ensures loss-less loading of cells to microfluidic platforms and can be easily adapted for downstream single cell analysis, e.g., to decode cellular interactions between different cell types.

This article presents a protocol for seeding scarce population of cells using pipette-tips to droplet microfluidic devices in order to provide higher encapsulation efficiency of cells in droplets.

一种基于管尖的液滴微流化平台种子细胞的方法

Amongst various microfluidic platform designs frequently used for cellular analysis, droplet-microfluidics provides a robust tool for isolating and ...

工程学

High Speed Droplet-based Delivery System for Passive Pumping in Microfluidic Devices

A novel microfluidic system has been developed that uses the phenomenon of passive pumping along with a user controlled droplet based fluid delivery system. Passive pumping is the phenomenon by which surface tension induced pressure differences drive fluid movement in closed channels. The automated fluid delivery system consists of a set of voltage controlled valves with micro-nozzles connected to a fluid reservoir and a control system. These voltage controlled valves offer a volumetrically precise way to deliver fluid droplets to the inlet of a microfluidic device in a high frequency manner. Based on the  dimensions demonstrated in the current study example, the system is capable of flowing 4 milliliters per minute (through a 2.2mm by 260um cross-sectional channel). Based on these same channel dimensions, fluid exchange of a point inside the channel can be achieved in as little as eight milliseconds. It is observed that there is interplay between momentum of the system (imparted by a combination of the droplets created by the valves and the fluid velocity in the channel), and the surface tension of the liquid. Where momentum provides velocity to the fluid flow (or vice-versa), equilibration of surface tension at the inlet provides a sudden stop to any flow. This sudden stop allows the user to control the flow characteristics of the channel and opens the door for a variety of biological applications, ranging anywhere from reagent delivery to drug-cell studies. It is also observed that when nozzles are aimed at the inlet at shallow angles, the droplet momentum can cause additional interesting fluid phenomena, such as mixing of multiple droplets in the inlet.

A novel microfluidic system has been developed using the phenomenon of passive pumping and a user controlled fluid delivery system. This microfluidic system has the potential to be used in a wide variety of biological applications given its low cost, ease of use, volumetric precision, high speed, repeatability and automation.

基于液滴在微流体装置的被动抽水高速输送系统

A novel microfluidic system has been developed that uses the phenomenon of passive pumping along with a user controlled droplet based fluid delivery ...

生物学

Digital Microfluidics for Automated Proteomic Processing

Clinical proteomics has emerged as an important new discipline, promising the discovery of biomarkers that will be useful for early diagnosis and prognosis of disease. While clinical proteomic methods vary widely, a common characteristic is the need for (i) extraction of proteins from extremely heterogeneous fluids (i.e. serum, whole blood, etc.) and (ii) extensive biochemical processing prior to analysis. Here, we report a new digital microfluidics (DMF) based method integrating several processing steps used in clinical proteomics. This includes protein extraction, resolubilization, reduction, alkylation and enzymatic digestion. Digital microfluidics is a microscale fluid-handling technique in which nanoliter-microliter sized droplets are manipulated on an open surface. Droplets are positioned on top of an array of electrodes that are coated by a dielectric layer - when an electrical potential is applied to the droplet, charges accumulate on either side of the dielectric. The charges serve as electrostatic handles that can be used to control droplet position, and by biasing a sequence of electrodes in series, droplets can be made to dispense, move, merge, mix, and split on the surface. Therefore, DMF is a natural fit for carrying rapid, sequential, multistep, miniaturized automated biochemical assays. This represents a significant advance over conventional methods (relying on manual pipetting or robots), and has the potential to be a useful new tool in clinical proteomics.
Mais J. Jebrail, Vivienne N. Luk, and Steve C. C. Shih contributed equally to this work.
Sergio L. S. Freire's current address is at the University of the Sciences in Philadelphia located at 600 South 43rd Street, Philadelphia, PA 19104.

Digital Microfluidics is a technique characterized by the manipulation of discrete droplets (~nL - mL) on an array of electrodes by the application of electrical fields. It is well-suited for carrying out rapid, sequential, miniaturized automated biochemical assays. Here, we report a platform capable of automating several proteomic processing steps.

数字微流体进行自动蛋白质组学处理

Clinical proteomics has emerged as an important new discipline, promising the discovery of biomarkers that will be useful for early diagnosis and ...

A Femtoliter Droplet Array for Massively Parallel Protein Synthesis from Single DNA Molecules

Advances in spatial resolution and detection sensitivity of scientific instrumentation make it possible to apply small reactors for biological and chemical research. To meet the demand for high-performance microreactors, we developed a femtoliter droplet array (FemDA) device and exemplified its application in massively parallel cell-free protein synthesis (CFPS) reactions. Over one million uniform droplets were readily generated within a finger-sized area using a two-step oil-sealing protocol. Every droplet was anchored in a femtoliter microchamber composed of a hydrophilic bottom and a hydrophobic sidewall. The hybrid hydrophilic-in-hydrophobic structure and the dedicated sealing oils and surfactants are crucial for stably retaining the femtoliter aqueous solution in the femtoliter space without evaporation loss. The femtoliter configuration and the simple structure of the FemDA device allowed minimal reagent consumption. The uniform dimension of the droplet reactors made large-scale quantitative and time-course measurements convincing and reliable. The FemDA technology correlated the protein yield of the CFPS reaction with the number of DNA molecules in each droplet. We streamlined the procedures about the microfabrication of the device, the formation of the femtoliter droplets, and the acquisition and analysis of the microscopic image data. The detailed protocol with the optimized low running cost makes the FemDA technology accessible to everyone who has standard cleanroom facilities and a conventional fluorescence microscope in their own place.

The overall goal of the protocol is to prepare over one million ordered, uniform, stable, and biocompatible femtoliter droplets on a 1 cm2 planar substrate that can be used for cell-free protein synthesis.

用于从单一DNA分子大规模平行蛋白质合成的Femtoliter滴体阵列

Advances in spatial resolution and detection sensitivity of scientific instrumentation make it possible to apply small reactors for biological and ...

化学

A Versatile Kit Based on Digital Microfluidics Droplet Actuation for Science Education

This paper describes an educational kit based on digital microfluidics. A protocol for luminol-based chemiluminescence experiment is reported as a specific example. It also has fluorescent imaging capability and closed humidified enclosure based on an ultrasonic atomizer to prevent evaporation. The kit can be assembled within a short period of time and with minimal training in electronics and soldering. The kit allows both undergraduate/graduate students and enthusiasts to obtain hands-on experience in microfluidics in an intuitive way and be trained to gain familiarity with digital microfluidics.

We describe an educational kit that allows users to execute multiple experiments and gain hands-on experience on digital microfluidics.

基于数字微流体的多功能套件，用于科学教育

This paper describes an educational kit based on digital microfluidics. A protocol for luminol-based chemiluminescence experiment is reported as a ...

Electrowetting-based Digital Microfluidics Platform for Automated Enzyme-linked Immunosorbent Assay

Electrowetting is the effect by which the contact angle of a droplet exposed to a surface charge is modified. Electrowetting-on-dielectric (EWOD) exploits the dielectric properties of thin insulator films to enhance the charge density and hence boost the electrowetting effect. The presence of charges results in an electrically induced spreading of the droplet which permits purposeful manipulation across a hydrophobic surface. Here, we demonstrate EWOD-based protocol for sample processing and detection of four categories of antigens, using an automated surface actuation platform, via two variations of an Enzyme-Linked Immunosorbent Assay (ELISA) methods. The ELISA is performed on magnetic beads with immobilized primary antibodies which can be selected to target a specific antigen. An antibody conjugated to HRP binds to the antigen and is mixed with H2O2/Luminol for quantification of the captured pathogens. Assay completion times of between 6 and 10 min were achieved, whilst minuscule volumes of reagents were utilized.

Electrowetting-based digital microfluidic is a technique that utilizes a voltage-driven change in the apparent contact angle of a microliter-volume droplet to facilitate its manipulation. Combining this with functionalized magnetic beads enables the integration of multiple laboratory unit operations for sample preparation and identification of pathogens using Enzyme-linked Immunosorbent Assay (ELISA).

基于电润的数字微流体平台，用于自动酶链接免疫吸附测定

Electrowetting is the effect by which the contact angle of a droplet exposed to a surface charge is modified. Electrowetting-on-dielectric (EWOD) exploits ...

用于组合插头生产的微流体装置的设置

Bilayer Microfluidic Device for Combinatorial Plug Production

Droplet microfluidics is a versatile tool that allows the execution of a large number of reactions in chemically distinct nanoliter compartments. Such systems have been used to encapsulate a variety of biochemical reactions - from incubation of single cells to implementation of PCR reactions, from genomics to chemical synthesis. Coupling the microfluidic channels with regulatory valves allows control over their opening and closing, thereby enabling the rapid production of large-scale combinatorial libraries consisting of a population of droplets with unique compositions. In this paper, protocols for the fabrication and operation of a pressure-driven, PDMS-based bilayer microfluidic device that can be utilized to generate combinatorial libraries of water-in-oil emulsions called plugs are presented. By incorporating software programs and microfluidic hardware, the flow of desired fluids in the device can be controlled and manipulated to generate combinatorial plug libraries and to control the composition and quantity of constituent plug populations. These protocols will expedite the process of generating combinatorial screens, particularly to study drug response in cells from cancer patient biopsies.

作者聚焦：在精准肿瘤学中整合计算和实验方法

The fabrication of a polydimethylsiloxane (PDMS)-based bilayer device for the production of combinatorial libraries in water-in-oil emulsions (plugs) is presented here. The necessary hardware and software required to automate plug production are detailed in the protocol, and the production of a quantitative library of fluorescent plugs is also demonstrated.

用于组合塞生产的双层微流控装置

Droplet microfluidics is a versatile tool that allows the execution of a large number of reactions in chemically distinct nanoliter compartments. Such ...

以减少液滴表面的相互作用来优化Bioanalytes运输中的数字微流体

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此视频中的章节

荧光检测方法，为微流体液滴平台

微流体芯片ICPMS进样

一种基于管尖的液滴微流化平台种子细胞的方法

基于液滴在微流体装置的被动抽水高速输送系统

数字微流体进行自动蛋白质组学处理

用于从单一DNA分子大规模平行蛋白质合成的Femtoliter滴体阵列

基于数字微流体的多功能套件，用于科学教育

基于电润的数字微流体平台，用于自动酶链接免疫吸附测定

用于组合塞生产的双层微流控装置

用于组合塞生产的双层微流控装置