私たちは、財政支援のためのLindback財団、博士アレクサンダー·シドレンコとエルザチュー実り議論や技術支援のために、そしてCと支援のための教授のロバート·スミスに感謝虫アッセイ 。

注：以下の手順では、実験室の安全ガイドラインは、必ず従わなければなりません。特に重要なのは、高電圧（&gt; 500 V）および取り扱い化学物質を扱う安全である。 キャンドルすすと導電性基板の1.コーティング<ol><li>長方形に切断の銅金属（0.5mm厚75×43ミリメートル）。約20秒間水道水で洗って、約30秒間銅エッチング液に浸漬することにより、各銅基板をきれいにし、紙で乾燥させます。 注：以下の方法1を使用する場合は、マシンに収まるように75×25 mmの寸法を変更します。 </li><li>ほぼ均一なすすコーティングを得るために、30〜45秒間、銅基板の下に点灯してパラフィンろうそくをスイープ（約40μm厚）。火炎内部〜1センチメートルで基板を保管してください。脆弱なすす面に触れないでください。 </li></ol> 2.コーティングでスート層の保護注：スート層は非常に脆弱である、および保護のために被覆しなければならない。つの単純な代替案（メソッド1及び2）は、ここで提案されているが、より堅牢なプロトコルが現在開発中である。 <ol><li>方法1 <ol><li>金属蒸発器またはスパッタリング装置内に試料をロードします。システムの操作手順に従って、チャンバを排気し、スート層（150〜200ナノメートル）の上に金の制御された堆積を開始します。デバイスは、室温まで冷却してみましょう。 </li><li>化学フード内部10分間1-ドデカンチオール溶液（1％v / v、95％エタノール中で、ACS / USPグレード）、ディップコート金属化基板。その後、60°に近い角度でデバイスを保持し、静かにエタノールのみの数滴で表面を洗う。一晩、デバイスは、乾燥してみましょう。 </li></ol></li><li>方法2 <ol><li>基板はまだろうそくの炎から温かいうちに化学フード内で、すぐに煤を基板上に塗布した後、のいずれかの側にフッ素化された液体の一部の液滴を堆積し基板、および90°に近い角度に基板を傾けます。より多くの液滴を堆積し、それらが全体のすす表面上を転がるましょう。 注：液滴がスポットに当たるときは、煤がその領域から洗い流される。フッ素化された液体の液滴の広がりを可能な限りう。 </li><li>化学フード内部のホットプレート（15分間160℃）上の基板焼く。 </li><li>基板は、使用前に室温で一晩放置します。無期限に保管してください。 </li></ol></li></ol> 3.上部電極の作製（Abdelgawad らから適応。12） <ol><li>グラフィックデザインソフトウェアを使用して電極を描画します。各電極は、幅0.3mm、長さ2mmであり、電極間のギャップは0.3mmである。接点（下記参照、コネクタにスナップするため）との間のギャップは2.3ミリメートルです（ 図1）。 </li><li>モナーク形式に柔軟な銅積層（厚さ35μm）（3.87×7.5インチ）トリム。他のサイズを使用してIプリンタと互換性fを。カラープリンタの手差しトレイに積層体をロードします。 </li><li>銅板に印刷する場合、エッチング中に印刷されたパターンを保護する、銅基板上のブラックインクの密度の高い層を可能にするために「リッチブラック」、または「登録黒&quot;を使用してください（詳細についてはAbdelgawad ら 12を参照）ことを確認してください。プリント基板に完全に乾燥させ、一晩。 </li><li>化学フード内では、（40°C）、銅エッチング液を50 mlのビーカーを温める。ビーカー内の印刷された積層体を浸し、ゆっくり約10分間溶液中にそれを振る。エッチング時間は、銅エッチング溶液によって異なります。数分ごとには、腐食を確認し、パターンが無傷であるかどうかを確認。 </li><li>慎重に水で積層体を洗って、化学フード内でアセトン、エタノールでコーティングを除去。もう一度洗って、優しくペーパータオルで積層体を乾燥させます。 </li><li>慎重にGLAに電極を積層板を取り付けるSSスライド（75×25ミリメートル、厚さの〜1ミリメートル）、両面テープを使用した。エアポケットを避けてください。 </li><li>テープを使用して、電極にパーフルオロアルコキシPFAのフィルムを取り付けます。これは、液滴と電極の偶発的な接触を防止するのに役立つによる短絡の損傷上部電極。 </li></ol> 4.（図2の回路）電子インターフェース<ol><li>ユニバーサル回路基板にリレーやコンデンサCを半田付けします。 </li><li>電子回路用のブレッドボード上で10のリレードライバの残りの部分を組み立てます。 </li><li>各ワイヤは、制御基板内のチャネルに、運転者の入力をリレー。 </li><li>慎重にコネクタ（ 図3）に上部電極をスナップ。図に示すように、ワイヤそれぞれは、上部電極にドライバの出力を中継する。電気ノイズを最小化するために、リレーからの一対のワイヤ間に接地されたコネクタ接点があることに留意されたい。 注：コネクターは、Tを制御するために調節可能なプラットフォーム上に座っ彼は距離頂部と底部との間（0.1〜0.5ミリメートル）（スートコーティングされた）基板。 </li><li>その後2345年、3456 など ; 0.8秒、作動させるために1234年、すなわち運動（の方向に1電極をずらし、同時に4電極に高電圧（HV）アプリケーション（約0.8秒）のタイミングを制御するプログラムを使用してください。0.8グループごとに秒、その後、後方に、反対方向に移動するので、液滴も同様）。 </li></ol> 5.液滴可視化と取扱い<ol><li> CCDカメラと組み合わせ96X倍率アセンブリ - 液滴の動きを記録するために、24Xで構成されている可視化システムを使用する。 Sビデオを使用してカメラにビデオレコーダーを接続します。 </li><li> Cを含む4μlの滴をピペットですす被覆基板の底部にあるメディアでエレガンス 。 </li><li>液滴上記〜0.3ミリメートルに上部電極を持参してください。液滴がより簡単に操作のための、ちょうど第五電極直下の、真ん中に近くなければなりません。 </li><li>電子インターフェースと高電圧（500 V RMS）の電源をオンにし、それが移動を開始するまでの液滴に上部電極の距離を調整する。上部電極は、液滴を接触させてはいけません。 </li><li>電気パルスに応答したデバイス上で成功した液滴転送回数を記録することによって、データを収集する。成功した実験は、少なくとも700滴転送、 すなわち 、各電気パルスの後1転送することを特徴とする。 </li><li>滴が5から10パルスに応答して、もはや動かないまで、継続的にデータを収集する。 注：表面が劣化し始めると、動きが近い液滴に上部電極にすることによって回復することができた。 </li></ol>

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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. 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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. 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The authors declare that they have no competing financial interests.

プロトコルの最も重要なステップは、直接、液滴を動かすの成功に関連付けられている、スート層の保護である。スート層（上記の方法1）を金属化することは、製造の成功の100％に近いことができる。しかし、最大動作時間は約10分である。おそらく、小滴のフラクションは、金属層の貫通孔煤湿潤される。フッ素化液体とのスート層をコーティングすることは、最も簡単かつ最速の代替で?...

液体を扱う装置の小型化は、「ラボオンチップ」のプラットフォームの開発のために最も重要である。この方向では、最後の二十年は、様々なアプリケーションで、マイクロフルイディクスの分野で大きな進歩を目撃した。1-5同封チャンネル（チャンネルマイクロフルイディクス）内の流体の輸送に対照的な、DMFは、電極のアレイ上の液滴を操作する。この技術の最も魅力的なメリットの一つは、流体を輸送するための可動部がないことで、動きを瞬時に電気信号をオフにすることによって停止される。 しかし、液滴の運動は、液滴の内容、ユニバーサル」ラボオンチップ」プラットフォームの確かに望ましくない特性に依存する。タンパク質や他の検体を含む液滴が移動不能になって、デバイス表面に固執する。間違いなく、これをDMFアプリケーションの適用範囲を広げるための主要な制限されている; 6-8不要な表面の汚れを最小にする選択肢は、潜在的に液滴コンテンツに影響を与える可能性が液滴またはその周囲に余分な化学種の添加を含む。 以前、我々のグループは、追加の添加剤（電界DW装置）なしで、DMF中の細胞およびタンパク質の輸送を可能にする装置を開発し9これは、液滴圧延を優先するデバイス·ジオメトリを有するキャンドルすす、10に基づいて、面を組み合わせることによって達成されたさらに、液滴表面相互作用を減少させる、液滴に上向きの力をもたらす。このアプローチでは、液滴の運動は、表面の濡れに関連付けられていない。11 以下に説明する詳細な方法の目的は、余分な添加剤を含まない、タンパク質、細胞、および生物全体を含む液滴を輸送することができるDMFデバイスを製造することである。フィールド-DWデバイスは、大部分が独立して液滴化学者の作業完全に制御プラットフォームへの道を開くRY。 ここでは、それを示す、本シミュレーションでは、デバイスの動作に必要な高電圧にもかかわらず、液滴の両端の電圧降下は、液滴内部bioanalytesにほとんど影響を示し、印加電圧のごく一部である。実際には、 線虫（C.エレガンス ）、生物学の研究のさまざまな目的で使用線虫を用いた予備試験では、電圧が印加されるとワームが乱さ泳ぐことを示している。

<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、我々は、DMF中のタンパク質の運動を可能にするためにフィールド-DWデバイスを使用している。特に、ウシ血清アルブミン（BSA）を有する液滴が以前に（添加剤なし）、他の著者によって報告されたものより2000倍高い濃度で移動することができる。これは滴と表面との間の相互作用の減少によるものであった; 図4は、蛍光標識したBSAを（。フレイレら参照実験?...

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.

デジタルマイクロフルイディクスにBioanalytesの輸送を最適化するための縮小液滴表面相互作用の活用

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 ビュー.  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.

ビデオ: 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.

マイクロ流体液滴のプラットフォームのための蛍光検出法

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

生物工学

Capillary-based Centrifugal Microfluidic Device for Size-controllable Formation of Monodisperse Microdroplets

Here, we demonstrate a simple method for the rapid production of size-controllable, monodisperse, W/O microdroplets using a capillary-based centrifugal microfluidic device. W/O microdroplets have recently been used in powerful methods that enable miniaturized chemical experiments. Therefore, developing a versatile method to yield monodisperse W/O microdroplets is needed. We have developed a method for generating monodisperse W/O microdroplets based on a capillary-based centrifugal axisymmetric co-flowing microfluidic device. We succeeded in controlling the size of microdroplets by adjusting the capillary orifice. Our method requires equipment that is easier-to-use than with other microfluidic techniques, requires only a small volume (0.1-1 &#181;l) of sample solution for encapsulation, and enables the production of hundreds of thousands number of W/O microdroplets per second. We expect this method will assist biological studies that require precious biological samples by conserving the volume of the samples for rapid quantitative analysis biochemical and biological studies.

Here, we demonstrate a simple production method for size-controllable, monodisperse, water-in-oil (W/O) microdroplets using a capillary-based centrifugal microfluidic device. This method requires only a small sample volume and enables high-yield production. We expect this method will be useful for rapid biochemical and cellular analyses.

単分散微小液滴のサイズ制御可能な形成のためのキャピラリーベースの遠心マイクロ流体デバイス

Here, we demonstrate a simple method for the rapid production of size-controllable, monodisperse, W/O microdroplets using a capillary-based centrifugal ...

工学

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.

ICPMSサンプル導入のためのマイクロ流体チップ

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.

ピペット チップを用いた液滴マイクロ流体プラットフォームに播種細胞メソッド

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

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.

マイクロ流体デバイスでパッシブポンピング用高速液滴ベースのデリバリーシステム

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.

自動プロテオーム処理のためのデジタルマイクロ流体

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.

単一DNA分子からの超並列タンパク質合成のためのフェムトリットルドロップレットアレイ

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.

科学教育のためのデジタルマイクロ流体ドロップレット作動に基づく多目的なキット

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).

酵素結合免疫吸着アッセイ用エレクトロウェッティングベースのデジタルマイクロ流体プラットフォーム

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

コンビナトリアルプラグ製造のためのマイクロ流体デバイスの構築

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.

著者スポットライト:Precision Oncologyにおける計算アプローチと実験的アプローチの統合

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.

コンビナトリアルプラグ製造のための二層マイクロ流体デバイス

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

デジタルマイクロフルイディクスにBioanalytesの輸送を最適化するための縮小液滴表面相互作用の活用

要約

さらに動画を探す

この動画の章

マイクロ流体液滴のプラットフォームのための蛍光検出法

単分散微小液滴のサイズ制御可能な形成のためのキャピラリーベースの遠心マイクロ流体デバイス

ICPMSサンプル導入のためのマイクロ流体チップ

ピペットチップを用いた液滴マイクロ流体プラットフォームに播種細胞メソッド

マイクロ流体デバイスでパッシブポンピング用高速液滴ベースのデリバリーシステム

自動プロテオーム処理のためのデジタルマイクロ流体

単一DNA分子からの超並列タンパク質合成のためのフェムトリットルドロップレットアレイ

科学教育のためのデジタルマイクロ流体ドロップレット作動に基づく多目的なキット

酵素結合免疫吸着アッセイ用エレクトロウェッティングベースのデジタルマイクロ流体プラットフォーム

コンビナトリアルプラグ製造のための二層マイクロ流体デバイス

デジタルマイクロフルイディクスにBioanalytesの輸送を最適化するための縮小液滴表面相互作用の活用

要約

さらに動画を探す

この動画の章

マイクロ流体液滴のプラットフォームのための蛍光検出法

単分散微小液滴のサイズ制御可能な形成のためのキャピラリーベースの遠心マイクロ流体デバイス

ICPMSサンプル導入のためのマイクロ流体チップ

ピペット チップを用いた液滴マイクロ流体プラットフォームに播種細胞メソッド

マイクロ流体デバイスでパッシブポンピング用高速液滴ベースのデリバリーシステム

自動プロテオーム処理のためのデジタルマイクロ流体

単一DNA分子からの超並列タンパク質合成のためのフェムトリットルドロップレットアレイ

科学教育のためのデジタルマイクロ流体ドロップレット作動に基づく多目的なキット

酵素結合免疫吸着アッセイ用エレクトロウェッティングベースのデジタルマイクロ流体プラットフォーム

コンビナトリアルプラグ製造のための二層マイクロ流体デバイス

ピペットチップを用いた液滴マイクロ流体プラットフォームに播種細胞メソッド