作者想感谢 DFG 的支持。

1. 磁性纳米粒子的水化 <ol> <li> 用于合成疏水性磁性微粒, 添加 0.85 g FeCl 3 六水合物 (3.14 摩尔) 和 0.30 g FeCl 2 四水合物 (1.51 摩尔) 至200毫升水/乙醇溶液 (4:1 伏/伏). </li>
	<li> 对此混合物, 增加0.20 毫升 1 h, 1 h, 2 h, 2 h Perfluorooctyltriethoxysilane (PFOTES) (5.23 摩尔) 与剧烈搅拌由磁性搅拌器 (500 rpm)。在惰性气体气氛中 (N 2 ) 进行合成, 使用带有软木塞的圆底烧瓶, 以防止磁铁矿微粒的二次氧化. </li>
	<li> 用氢氧化铵溶液 (1.5 米) 将溶液下落调整到 ph 值 8 (ph 值由 ph 计测定)。使用磁力搅拌器将溶液搅拌24小时. </li>
	<li> 通过将烧瓶放置在条形磁铁上 (钕长方体磁铁 40 x 20 x 10 mm, 粘附力25千克) 将颗粒从溶液中磁性分离出来。倒出的解决方案, 同时保持磁铁的底部的瓶子. </li>
	<li> 用水/乙醇溶液清洗粒子三次, 而使用的是1.4 中描述的条形磁铁)。在60和 #176 处将颗粒干燥; C 为 24 h (产生 ca 0.43 g). </li>
	<li> 用于分析粒子, 根据制造商和 #39 的说明使用扫描电子显微镜. </li>
</ol> 2。反应器的制作 <ol> <li> 使用玻璃杵将所有干燥的微粒磨碎, 然后将它们直接放入称重锅 (46 和 #215; 46 和 #215; 8 mm, 聚苯乙烯) </li>
	<li> 吸管10和 #181; l 反应溶液 (如5.1 所述的成分) 到所有的粒子上, 并在大约十年代轻微地移动称重盘 (粒子质量为10和 #181; 微: ca 3.2 x 10 -7 kg类 = "xref" > 33 )。在室温下储存剩余的微粒 (在反应溶液周围没有组装的微粒) 以供进一步应用. </li>
	<li> 用于测量微的接触角, 请建立一个5和 #181; 微 2.2), 将其放置在聚四氟乙烯薄膜上, 并根据制造商和 #39 使用光学接触角测量装置分析接触角;说明. </li>
</ol> 3. 3 d-打印线圈体 <ol> <li> 设计双线圈体, 其高度为 16 mm (一室), 直径为 10 mm, 内径为 ca. 4 mm, 并根据制造商和 #39 使用 CAD 软件;说明. </li>
	<li> 根据制造商和 #39 的指示, 使用聚乳酸灯丝等材料打印带3D 打印机的线圈体。用一台电脑控制的绕线机将车身用0.08 毫米铜线包裹起来, 实现4500绕组. </li>
</ol> 4。制造的驱动平台 <ol> <li> 在一个矩阵 ( 如 3 x 3 矩阵) 上安排双线圈, 在一个带有珀尔卡元件的电路板上, 拧紧双线圈, 然后通过带状电缆将它们连接到一个控件 ( 图 3 ). </li>
	<li> 根据所需的应用, 添加一个铁芯 (32 毫米高, 4 毫米直径) 或钕磁铁 (12.5 毫米高度, 4 毫米直径, 1035 kA m -1 ), 以获得更强的磁场. </li>
	<li> 完成平台放置一个板, 最好是石英玻璃, 最大高度为 1 mm 的线圈矩阵. </li>
	<li> 在平台表面放置一个微. </li>
	<li> 从反应溶液中提取上部粒子, 从而打开一个微, 使用4.1 中提到的控制来激活内部的钕磁铁。关闭微再次停用线圈. </li>
	<li> 合并两个初始的反应器 ca. 10 mm 使用钕磁体, 如4.2 所述)。通过激活 ca 的线圈, 在所需的线圈体中向上提起磁体. 二十五年代打开一个微 (在微和磁铁之间所需的距离约 12 mm), 并将另一个移动到平台上的相同位置. </li>
	<li> 在平台表面的微上冷却反应液, 并减少在线圈矩阵下方的贴珀元件上的线圈温度开关, 如4.1 所述). </li>
</ol> 5。通过合并反应器的酶反应 <ol> <li> 将辣根过氧化物酶溶解在浓度为0.1 和 #181; g mL -1 在磷酸钾缓冲 (0.1 M, pH 6.5)。稀释基体, 10-乙酰基 37-dihydroxyphenoxazine (10 毫米在砜 (亚砜)) 与磷酸钾缓冲 (0.1 M, pH 6.5) 的浓度200和 #181; m. </li>
	<li> 使用10和 #181; 每个解决方案中的 L 分别构建两个反应器, 如2.2 所述)。将两个反应器用磁力的平均值 (钕圆柱磁铁: 12.5 mm x 4 mm, 1035 kA m -1 ) 合并为4.6 和 #176; C. </li>
	<li> 通过定位荧光探针检测反应 (激发波长: 570 nm, 发射波长: 585 nm) ca 10 mm 直接在合并之前打开的微. </li>
</ol>

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作者没有什么可透露的。

为了成功地利用微流控技术, 必须将反应量与生物科技合成和分析的要求相对应。在这里提出的驱动平台, 可以移动微流控液滴的磁力。通过将液滴与磁性疏壳封闭在反应平台的平面表面上, 可以在两个维度上自由地进行运动。因此, 在传统的微流控系统中, 采用具有复杂流量控制装置的预定义微流控通道系统来传递溶剂和样品。因此, 小型反应液滴的自动驱动是对已知的实验室芯片平台的显著简化?...

微型反应的技术应用主要是在预定义微通道芯片中进行的。这些系统在文献中得到广泛的建立和全面的描述 (特别是1,2,3)。在 2011年, 微流控技术的营业额在全球总计62亿欧元4。相比之下, 自由移动的微型反应堆舱室的使用以前只在有限的范围内进行过审查和出版。最常见的移动水下微滴的方法是润湿5。其他方法为下落在表面上是基于电场6, 磁力力量7或声学驱动8。由于其不利的表面体积比, 这些 droplet-based 微系统暴露在强烈的蒸发效应。因此, 下落运动通常建立作为液体 two-phase 系统, 其中上部阶段有一个高沸点保护水相从蒸发。然而, 这种方法有很大的风险, 污染的反应液滴不受控制的扩散。这是对上述系统的技术建立的一个重大障碍。
最近的工作涉及换药液固相转变。一种高效的方法是使用疏表面, 从而形成球形水滴。这个反应概念的引伸是使用微反应隔间与疏表面或壳, 例如可能包括聚四氟乙烯 (PTFE) 微粒9。他们的接触角度在表面上通常是在160范围内 (取决于表面粗糙度)。因此, 球形隔间提供了对表面运动的最小阻力, 同时提供了防止水蒸发的保护措施。
含微 PTFE 颗粒的水性水滴可以保持其球形形状, 直径可达2毫米左右。在更高的容量, 疏水壳通常不是完全地闭合的10。利用离子液体12, 对其它壳体材料的影响以及液体大理石在非极性溶剂中的应用范围进行了研究。为形成疏水性基于壳, 到目前为止微粒直径在大小的10毫微米-30 µm 被描述了11,14,16。新的研究表明, 疏水性纳米粒子作为壳体材料, 比微粒13的使用更为有效。第一个稳定性研究证实, 当粒子大小从 ca 减少时, 稳定性提高了.600 nm 到ca 100 nm。这可能是由于在水球体周围的密度更大的粒子分布15。
水的反应隔间的保护由疏水壳和他们的指定作为液体大理石在2001年首先被描述了由 Aussillous et al.和 Mahadevan et al.17,18. 从那以后, 这些定义的反应隔间的应用很少被描述。例如, 基于液体弹珠19的气体传感器和基于光学定性基础的水污染检测方法已被开发成20。作者区分了高反应率和低消耗化学物质的微反应系统的优势。最近的出版物涉及生产 pH 敏感的液体弹珠16或表示 "两面" 粒子与两种不同的功能涂层。例如, Bormashenko et al.可以合成一个微, 其外壳由聚四氟乙烯和半导体炭黑21组成。此外, 通过渗透气液界面24, 反应器能高效、方便地吸收外氧作为单体的合成 polyperoxides。在另一种方法中, 二氧化硅-基于液弹的壳体提供反应基板表面来调节经典的银镜反应26。亲水-核-疏水壳液滴的研究和开发目前存在的问题是粒度调整、单分散液滴的可再生生产、表面润湿性和第二次的效果亲水壳在微反应舱室22, 以及更好地控制液滴轨迹,例如用于开发连续 microPCR 系统4。
这些反应器的磁性驱动提供了相对地高运动范围的好处和一个好选择力量, 当工作在生物化学的系统时。在使用疏水磁铁矿粒子时, 它们既实现了磁力传递到反应器运动的功能, 也满足了疏水壳的作用。在 2006年, 莱曼et al.第一次被假定为液滴内磁性微粒的磁性运动。23和田et al.25, 谁使用手动移动永久磁铁作为执行器的一个单一的水滴的动员。另一种移动少量液体的方法是由赵et al.实现的, 后者使用疏水 Fe3O4粒子作为磁壳。磁性液体大理石的壳在下落的上部被打开了由垂直的反向磁场27。基于这一概念, 薛et al.能够开发出形成微的粒子, 其表面张力为20.1 达因 cm− 1 28。Lin et al.捏造了新的纤维素微/纳米分层球体与 superparamagnetism 和棉织, 提供了神的稳定性磁液滴运输和操作31。到目前为止, 这只是作为一个原理的研究, 而不是用于任何应用程序。液体弹珠的磁性和电控目前在第一种方法中进行。赵et al.在 2010年15和张et al. 2012 29中, 可以通过手动操作在核心壳液滴下的永磁体运动来进行液滴操作。Bormashenko et al.11通过接近钕磁铁, 实现了将铁磁性液体大理石的加速度提高到 25 cm s-1的速度。上述原则研究完全是由一个小型永磁体的手动运动进行的。作为下一步的发展步骤, 赵et al.最近能够通过改变永磁体的距离来估计磁性液体大理石运动所需的磁通密度30。对于与普通实验室芯片系统相媲美的反应控制, 提供自动控制离散液体 v 的方法似乎是不可避免的。olumes为了满足这一需要, 我们开发了一种新的基于变场梯度的控制系统, 以固定、移动和打开磁反应器。

<table><tbody><tr><td>3D-printer</td><td>FelixPrinters</td><td>Pro1</td><td></td></tr><tr><td>10-acetyl-3,7-dihydroxyphenoxazine (Amplex Red)</td><td>Life Technologies</td><td>A12222</td><td></td></tr><tr><td>Ammonium hydroxide</td><td>TU-KL</td><td>1072</td><td></td></tr><tr><td>CAD software</td><td>Siemens</td><td>Soled edge</td><td></td></tr><tr><td>Contact angle measuring device</td><td>Dataphysics</td><td>OCA 20</td><td></td></tr><tr><td>Cylinder magnet</td><td>Webcraft GmbH</td><td>S-04-13-N</td><td>https://www.supermagnete.de/stabmagnete-neodym-rund/stabmagnet-durchmesser-4mm-hoehe-12.5mm-neodym-n42-vernickelt_S-04-13-N</td></tr><tr><td>Dipotassium phosphate</td><td>Bernd Kraft</td><td>7758-11</td><td></td></tr><tr><td>Drying oven</td><td>Binder</td><td>FD 115</td><td></td></tr><tr><td>Ethanol</td><td>Sigma-Aldrich</td><td>68-17-5</td><td></td></tr><tr><td>FeCl&lt;sub&gt;2&lt;/sub&gt; tetrahydrate</td><td>TU-KL</td><td>1625</td><td></td></tr><tr><td>FeCl&lt;sub&gt;3&lt;/sub&gt; hexahydrate</td><td>TU-KL</td><td>1622</td><td></td></tr><tr><td>Fluorescence probe</td><td>PerkinElmer</td><td>LS 55</td><td></td></tr><tr><td>Horseradish peroxidase</td><td>Carl Roth</td><td>9003-99-0</td><td></td></tr><tr><td>Hydrogen peroxide</td><td>Th.Geyer GmbH &amp;amp; Co</td><td>7722-84-1</td><td></td></tr><tr><td>Monopotassium phosphate</td><td>Bernd Kraft</td><td>7778-77-0</td><td></td></tr><tr><td>Peltier element</td><td>Conrad&amp;nbsp;</td><td>193569</td><td></td></tr><tr><td>Perfluoroctyltriethoxysilane</td><td>Sigma-Aldrich</td><td>51851-37-7</td><td></td></tr><tr><td>Scanning Electron Microscope</td><td>FEI</td><td>Helios NanoLab 650 DualBeam</td><td></td></tr><tr><td>Separation bar magnet</td><td>Webcraft GmbH</td><td>Q-40-20-10-N</td><td></td></tr><tr><td>Winding machine</td><td>IWT GmbH</td><td>FW122</td><td></td></tr></tbody></table>

aqueous droplets used as enzymatic microreactors and their electromagnetic actuation

为了成功地实现微流体反应系统, 如 PCR 和电泳, 小液体体积的运动是必不可少的。在传统的实验室芯片平台中, 溶剂和样品通过具有复杂流量控制装置的微流控通道传递。在这里提出的液滴驱动平台是一个有前途的选择。有了它, 它是可能移动液体下落 (微) 在一个反应平台的平面表面 (lab-in-a-下落)。反应器在平台疏水表面的驱动是基于对液滴外层的磁力作用, 它是由一层薄薄的疏磁铁矿颗粒制成的。平台的疏水表面是需要的, 以避免任何接触之间的液体核心和表面, 使微的平滑移动。在平台上, 可以同时定位和移动一个或多个带有10µL 卷的反应器。该平台本身包括一个 3 x 3 矩阵的电气双线圈, 其中既容纳钕或铁芯。磁场梯度是自动控制的。通过磁场梯度的变化, 可以自动操纵反应器的磁性疏水壳, 以使微或打开壳体可逆地运动。基质和相应的酶的反应可以通过合并反应器或使它们与表面固定化催化剂接触来启动。

贝壳颗粒的直径约640纳米。在这个 fluorosilane 壳颗粒内的磁化纳米粒子的直径在22纳米和 37 nm 之间。一个5µL 微与水作为一个液体核心有一个接触角的周围160。
如上所述, 移动10µL 微所需的力是1.34 ±0.08 µN.图 1显示了铜线线圈的电磁力和4500毫安驱动的内铁芯。通过有限元模型 (FEM) 估计力分布, 以确定所需?...

Watch this Scientific Journal Video about Aqueous Droplets Used as Enzymatic Microreactors and Their Electromagnetic Actuation at JoVE.com

Aqueous Droplets Used as Enzymatic Microreactors and Their Electromagnetic Actuation

Lab-in-a 滴反应系统允许在微流控尺度上实现复杂反应的多用途。一个由 3 x 3 电磁线圈组成的自动驱动平台被开发并成功地用于合并两个10µL 反应器, 从而在产生的液体弹珠中启动酶反应。

用作酶反应器的水滴及其电磁驱动

For the successful implementation of microfluidic reaction systems, such as PCR and electrophoresis, the movement of small liquid volumes is essential. In ...

aqueous-droplets-used-as-enzymatic-microreactors-their

Research

JoVE Journal

Bioengineering

5.3K 次观看.  University of Kaiserslautern. Lab-in-a 滴反应系统允许在微流控尺度上实现复杂反应的多用途。一个由 3 x 3 电磁线圈组成的自动驱动平台被开发并成功地用于合并两个10µL 反应器, 从而在产生的液体弹珠中启动酶反应。

视频: Aqueous Droplets Used as Enzymatic Microreactors and Their Electromagnetic Actuation

The Visual Colorimetric Detection of Multi-nucleotide Polymorphisms on a Pneumatic Droplet Manipulation Platform

A simple and visual method to detect multi-nucleotide polymorphism (MNP) was performed on a pneumatic droplet manipulation platform on an open surface. This approach to colorimetric DNA detection was based on the hybridization-mediated growth of gold nanoparticle probes (AuNP probes). The growth size and configuration of the AuNP are dominated by the number of DNA samples hybridized with the probes. Based on the specific size- and shape-dependent optical properties of the nanoparticles, the number of mismatches in a sample DNA fragment to the probes is able to be discriminated. The tests were conducted via droplets containing reagents and DNA samples respectively, and were transported and mixed on the pneumatic platform with the controlled pneumatic suction of the flexible PDMS-based superhydrophobic membrane. Droplets can be delivered simultaneously and precisely on an open-surface on the proposed pneumatic platform that is highly biocompatible with no side effect of DNA samples inside the droplets. Combining the two proposed methods, the multi-nucleotide polymorphism can be detected at sight on the pneumatic droplet manipulation platform; no additional instrument is required. The procedure from installing the droplets on the platform to the final result takes less than 5 min, much less than with existing methods. Moreover, this combined MNP detection approach requires a sample volume of only 10 &#181;l in each operation, which is remarkably less than that of a macro system.

This work presents a simple and visual method to detect multi-nucleotide polymorphisms on a pneumatic droplet manipulation platform. With the proposed method, the entire experiment, including droplet manipulation and detection of multi-nucleotide polymorphisms, can be performed near 23 &#176;C without the aid of advanced instruments.

在气动操纵液滴平台多核苷酸多态性的视觉比色法检测

A simple and visual method to detect multi-nucleotide polymorphism (MNP) was performed on a pneumatic droplet manipulation platform on an open surface. ...

科研

遗传学

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 ...

生物工程

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.

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; ...

工程学

A Versatile Automated Platform for Micro-scale Cell Stimulation Experiments

Study of cells in culture (in vitro analysis) has provided important insight into complex biological systems. Conventional methods and equipment for in vitro analysis are well suited to study of large numbers of cells (&ge;105) in milliliter-scale volumes (&ge;0.1 ml). However, there are many instances in which it is necessary or desirable to scale down culture size to reduce consumption of the cells of interest and/or reagents required for their culture, stimulation, or processing. Unfortunately, conventional approaches do not support precise and reproducible manipulation of micro-scale cultures, and the microfluidics-based automated systems currently available are too complex and specialized for routine use by most laboratories. To address this problem, we have developed a simple and versatile technology platform for automated culture, stimulation, and recovery of small populations of cells (100 - 2,000 cells) in micro-scale volumes (1 - 20 &mu;l). The platform consists of a set of fibronectin-coated microcapillaries ("cell perfusion chambers"), within which micro-scale cultures are established, maintained, and stimulated; a digital microfluidics (DMF) device outfitted with "transfer" microcapillaries ("central hub"), which routes cells and reagents to and from the perfusion chambers; a high-precision syringe pump, which powers transport of materials between the perfusion chambers and the central hub; and an electronic interface that provides control over transport of materials, which is coordinated and automated via pre-determined scripts. As an example, we used the platform to facilitate study of transcriptional responses elicited in immune cells upon challenge with bacteria. Use of the platform enabled us to reduce consumption of cells and reagents, minimize experiment-to-experiment variability, and re-direct hands-on labor. Given the advantages that it confers, as well as its accessibility and versatility, our platform should find use in a wide variety of laboratories and applications, and prove especially useful in facilitating analysis of cells and stimuli that are available in only limited quantities.

We have developed an automated cell culture and interrogation platform for micro-scale cell stimulation experiments. The platform offers simple, versatile, and precise control in cultivating and stimulating small populations of cells, and recovering lysates for molecular analyses. The platform is well suited to studies that use precious cells and/or reagents.

一种多功能自动化平台微尺度细胞刺激实验

Study of cells in culture (in vitro analysis) has provided important insight into complex biological systems. Conventional methods and equipment for in ...

Microfluidic Preparation of Liquid Crystalline Elastomer Actuators

This paper focuses on the microfluidic process (and its parameters) to prepare actuating particles from liquid crystalline elastomers. The preparation usually consists in the formation of droplets containing low molar mass liquid crystals at elevated temperatures. Subsequently, these particle precursors are oriented in the flow field of the capillary and solidified by a crosslinking polymerization, which produces the final actuating particles. The optimization of the process is necessary to obtain the actuating particles and the proper variation of the process parameters (temperature and flow rate) and allows variations of size and shape (from oblate to strongly prolate morphologies) as well as the magnitude of actuation. In addition, it is possible to vary the type of actuation from elongation to contraction depending on the director profile induced to the droplets during the flow in the capillary, which again depends on the microfluidic process and its parameters. Furthermore, particles of more complex shapes, like core-shell structures or Janus particles, can be prepared by adjusting the setup. By the variation of the chemical structure and the mode of crosslinking (solidification) of the liquid crystalline elastomer, it is also possible to prepare actuating particles triggered by heat or UV-vis irradiation.

This article describes the microfluidic process and parameters to prepare actuating particles from liquid crystalline elastomers. This process allows the preparation of actuating particles and the variation of their size and shape (from oblate to strongly prolate, core-shell, and Janus morphologies) as well as the magnitude of actuation.

液晶弹性体致动器的微流控制备

This paper focuses on the microfluidic process (and its parameters) to prepare actuating particles from liquid crystalline elastomers. The preparation ...

化学

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 ...

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 ...

Glass-Based Devices to Generate Drops and Emulsions

In this manuscript, three different step-by-step protocols to generate highly monodisperse emulsion drops using glass-based microfluidics are described. The first device is built for the generation of simple drops driven by gravity. The second device is designed to generate emulsion drops in a coflowing scheme. The third device is an extension of the coflowing device with the addition of a third liquid that acts as an electric ground, allowing the formation of electrified drops that subsequently discharge. In this setup, two of the three liquids have an appreciable electrical conductivity. The third liquid mediates between these two and is a dielectric. A&#160;voltage difference applied between the two conducting liquids creates an electric field that couples with hydrodynamic stresses of the coflowing liquids, affecting the jet and drop formation process. The addition of the electric field provides a path to generate smaller drops than in simple coflow devices and for generating particles and fibers with a wide range of sizes.

Here, a protocol to manufacture glass-based microfluidic devices used for generating highly monodisperse emulsions with controlled drop size is presented.

基于玻璃的设备，用于产生液滴和乳液

In this manuscript, three different step-by-step protocols to generate highly monodisperse emulsion drops using glass-based microfluidics are described. ...

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 ...

Automated Microbial Cultivation and Adaptive Evolution using Microbial Microdroplet Culture System (MMC)

Conventional microbial cultivation methods usually have cumbersome operations, low throughput, low efficiency, and large consumption of labor and reagents. Moreover, microplate-based high-throughput cultivation methods developed in recent years have poor microbial growth status and experiment parallelization because of their low dissolved oxygen, poor mixture, and severe evaporation and thermal effect. Due to many advantages of micro-droplets, such as small volume, high throughput, and strong controllability, the droplet-based microfluidic technology can overcome these problems, which has been used in many kinds of research of high-throughput microbial cultivation, screening, and evolution. However, most prior studies remain&#160;at the stage of laboratory construction and application. Some key issues, such as high operational requirements, high construction difficulty, and lack of automated integration technology, restrict the wide application of droplet microfluidic technology in microbial research. Here, an automated Microbial Microdroplet Culture system (MMC) was successfully developed based on droplet microfluidic technology, achieving the integration of functions such as inoculation, cultivation, online monitoring, sub-cultivation, sorting, and sampling required by the process of microbial droplet cultivation. In this protocol, wild-type Escherichia coli (E. coli) MG1655 and a methanol-essential E. coli strain (MeSV2.2) were taken as examples to introduce how to use the MMC to conduct automated and relatively high-throughput microbial cultivation and adaptive evolution in detail. This method is easy to operate, consumes less labor and reagents, and has high experimental throughput and good data parallelity, which has great advantages compared with conventional cultivation methods. It provides a low-cost, operation-friendly, and result-reliable experimental platform for scientific researchers to conduct related microbial research.

Automated Microbial Cultivation and Adaptive Evolution using Microbial Microdroplet Culture System MMC

This protocol describes how to use the Microbial Microdroplet Culture system (MMC) to conduct automated microbial cultivation and adaptive evolution. MMC can cultivate and sub-cultivate microorganisms automatically and continuously and monitor online their growth with relatively high throughput and good parallelization, reducing labor and reagent consumption.