著者は、財政支援のためにロバート·W·ドイツ財団、防衛脅威削減局（DTRA）、研究とイノベーションの国立科学財団新興フロンティア（EFRI）を認める。著者はまた、クリーンルーム施設のサポートのためにメリーランド州NanocenterとそのFablabに感謝します。

マイクロ流体チップの微細加工1。 <ol><li>電気化学チップを準備する<ol><li>柄ゴールド電極<ol><li>アセトン、メタノール、及びイソプロパノール（「AMI」クリーン）ブランク4インチシリコンウェハ（プライムグレードの品質）をすすぐ。 N 2銃で乾燥させ、脱イオン水（DI）で、ウェハからイソプロパノールを洗い流す。 </li><li>プラズマ強化化学蒸着（PECVD）ツールを使用して厚さ1μmのSiO 2パッシベーション層を成長させる。 DCスパッタツールでの金の2000オングストロームの厚さの層に続いて、クロムの200オングストロームの厚さの層を堆積させる。 </li><li>スピン &quot;PR1&quot;ポジ型フォトレジスト（スピニングパラメータ：3000回転、1000回転/秒、30秒）は、1.6μmの厚さの均一なフィルム〜を作成します。ホットプレート上で100℃で1分間、ウェーハプリベーク。 </li><li> 190 MJ /フォトマスクを介して405nmの波長でのcm 2のウェハを公開します。 352 devに30秒間ウェハを開発するDIを有するウェハおよびN 2銃で乾燥をeloperとすすぎます。 </li><li> 1.5分間の金エッチング液で金をエッチングする。 〜30秒間クロムエッチング液でクロムをエッチングする。 DIを有するウェーハをすすぎ、N 2銃で乾燥。 </li><li> 「AMI」きれいな手順でフォトレジストを除去。 注：強い酸化剤および潜在的に爆発。 </li><li> 1 H 2 SO 4：4：4でウェハをクリーニングし、H 2 O 2混合物（ &quot;ピラニア&quot;クリーン）を1分間。 DIを有するウェーハをすすぎ、N 2銃で乾燥。 </li></ol></li><li>パターンプラチナ電極<ol><li>スピン「PR2」ポジ型フォトレジスト（スピニングパラメータ：3000回転、1000回転/秒、30秒）は、1.4ミクロンの厚さの均一なフィルム〜を作成します。ホットプレート上で100℃で1分間、ウェーハプリベーク。 </li><li>フォトマスクを介して405nmの波長では30mJ / cm 2のウエハを露光する。ホットプレート上で125℃で45秒間ウェハを焼く。洪水公開するを1000mJ /フォトマスクなしで405nmの波長でのcm 2のウエハ。 6の2分間、ウェーハを開発する：1 AZ400K開発者およびDIを有するウェーハをリンスし、N 2銃で乾燥。 </li><li>電子ビーム蒸着装置を用いて、白金を1,600オングストロームの厚さの層に続いてチタンの400オングストロームの厚さの層を堆積させる。 </li><li>アセトンとイソプロパノールですすぎ、続いて5分間超音波アセトン浴中にフォトレジストをオフに持ち上げます。 DIを有するウェーハをすすぎ、N 2銃で乾燥。 </li></ol></li></ol></li><li>アッセイチャネルを準備<ol><li>アッセイチャネル用の金型を準備する<ol><li> 「AMI」きれいな手順で空白の4インチシリコンウェハ（テストグレードの品質）をすすぐ。 DIとウェハからイソプロパノールをすすぎ、N 2銃で乾燥させた。 </li><li>スピン「PR3」ネガ型フォトレジスト（2段階の回転パラメータ：ステップ1 - 600回転、120回転/秒、10秒2のステップ - 1150 rpmで383回転/秒、〜100μmの厚さの均一な膜を作成するための27秒）。ホットプレート上のウエハ（2段階のベーキングパラメータプリベーク：ステップ1 - 300℃/ hrで65℃まで室温から立ち上げ、10分間温度を保持するステップ2 - 95℃まで上昇します。 300℃/時間で30分間温度を保持する）。 </li><li> 2,500 MJ /フォトマスクを介して405nmの波長でのcm 2のウェハを公開します。ポストベークウェハ300℃/ hrで95℃まで上昇させると、ホットプレート上で10分間温度を保持することによって。プロピレングリコールモノメチルエーテルアセテート（PGMEA）、開発中で10分間、ウェーハを開発する。イソプロパノールでウェーハを洗浄し、そしてN 2銃で乾燥。 </li></ol></li><li>アッセイチャネルを製作<ol><li> 20分間の真空チャンバ内に1ポリジメチルシロキサン（PDMS）混合物（エラストマーを20g、硬化剤の2g）および完全に脱ガス：10を準備する。 </li><li>アルミホイルでモールドウエハの周囲のエンクロージャを定義して、ゆっくりと未硬化PDMSを注ぐ金型の上。 </li><li>箱型炉内のPDMSを有する型焼く（2段階のベーキングパラメータ：ステップ1 - 5分で80℃、室温からランプアップステップ2 - 17分間80℃で保持）。 </li><li>アルミ箔上のカビや場所からピール離れてPDMS。外科医ナイフでアッセイチップをカットし、生検パンチツールで穴を定義します。 </li></ol></li></ol></li></ol> 2デバイスを組み立てる<ol><li>手動での電気化学チップ上の作用電極とPDMSアッセイチャンネルを合わせます。 PDMSはチャネルを密閉し、漏出を防ぐ、電気化学チップによく付着する。 </li><li>プラスチック肘やストレートコネクタアダプタ（OD 0.1875 &#39;とID 0.0625&#39; &#39;）として、短いフレキシブルチューブを使用してフレキシブルチューブ（外径0.087&#39;とID 0.015を &#39;&#39;）を接続します。デバイスの穴パンチの入口のそれぞれにコネクタを接続します。 </li><li>のもう一方の端にシリンジを接続しますチューブとシリンジポンプにシリンジを配置します。交互にセクション3の必要な手順のステップとそれに対応する解決策に従った注射器のセット、チューブ、コネクタを変更してください。 </li></ol> 3 DNAハイブリダイゼーションを分析注：チャネル全体が満たされるまでに200μl/時の流量で流路にゆっくりと各ソリューションを紹介。 <ol><li>別の一本鎖DNAプローブ配列を持つ各垂直マイクロチャネルを機能化（3つの別個の垂直マイクロチャンネルを並列に実行します）： <ol><li>それぞれ異なるssDNAのプローブインキュベーション溶液でマイクロチャネルを埋める（10 mMリン酸緩衝液、100mMのNaCl、10μMのトリス（2 - カルボキシエチル）ホスフィン（TCEP）、および1μMのプローブのssDNA）、およびマイクロチャネルをすすぎ、続いて1時間インキュベートPBSで。 </li><li> 10mMのPBS緩衝液中に6 - メルカプト-1 - ヘキサノール（MCH）の1mMのを含む溶液でマイクロチャネルを充填し、100mMのNaClおよび1.395 mMのTCEP、及びincuPBSでマイクロチャネルをリンスし1時間譲り。 </li></ol></li><li> 、PDMSを持ち上げ水平方向に90°回転し、独特のカウンターと基準電極（図S1）を含む各行に反応室の個別の行を公開するダウン置く。 </li><li> 4倍の対照測定溶液でマイクロチャネルを充填し、生理食塩水クエン酸ナトリウム（SSC）緩衝液を濃縮し、20分間インキュベートする。 </li><li>バックグラウンド測定は;（作業、カウンタに接続されたポテンショスタットを用いて、参照電極を5mMのPBS溶液中/ 5mMのフェロシアン化をフェリシアンでマイクロチャネルを満たし、（EIS電気化学インピーダンス分光法）、異なる周波数のための電気化学システムのインピーダンス値を記録EISパラメータ：1メガヘルツから0.1ヘルツ、年間あたり10周波数データ点、25 mVの振幅、参照電極に対して5mVの偏光）の周波数範囲。マイクロチャネル内の各作用電極のために、この測定を繰り返します。 </李&gt; <li> （各非相補相補鎖DNAのターゲットに対して実行する）DNAハイブリダイゼーションを測定します。 <ol><li> 4倍の目標のssDNA測定溶液とマイクロチャンネルを満たし、目標のssDNAの1μMを含むSSC緩衝液を濃縮し、20分間インキュベートする。 3.4段階に記載のDNAを測定します。 </li></ol></li></ol>

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Bioelectron. 38, 114-120 (2012).</li><li>Wink, T., van Zuilen, S. J., Bult, A., van Bennekom, W. P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Self-assembled+monolayers+for+biosensors.">Self-assembled monolayers for biosensors.</a> Analyst. 122, 43R-50R (1997).</li><li>McEwen, G. D., Chen, F., Zhou, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Immobilization,+hybridization,+and+oxidation+of+synthetic+DNA+on+gold+surface:+Electron+transfer+investigated+by+electrochemistry+and+scanning+tunneling+microscopy.">Immobilization, hybridization, and oxidation of synthetic DNA on gold surface: Electron transfer investigated by electrochemistry and scanning tunneling microscopy.</a> Anal. Chim. Acta. 643, 26-37 (2009).</li><li>Arinaga, K., Rant, U., Tornow, M., Fujita, S., Abstreiter, G., Yokoyama, N. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+role+of+surface+charging+during+the+coadsorption+of+mercaptohexanol+to+DNA+layers+on+gold:+Direct+observation+of+desorption+and+layer+reorientation.">The role of surface charging during the coadsorption of mercaptohexanol to DNA layers on gold: Direct observation of desorption and layer reorientation.</a> Langmuir. 22, 5560-5562 (2006).</li><li>Katz, E., Willner, I. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Probing+biomolecular+interactions+at+conductive+and+semiconductive+surfaces+by+impedance+spectroscopy:+Routes+to+impedimetric+immunosensors,+DNA-sensors,+and+enzyme+biosensors.">Probing biomolecular interactions at conductive and semiconductive surfaces by impedance spectroscopy: Routes to impedimetric immunosensors, DNA-sensors, and enzyme biosensors.</a> Electroanalysis. 15, 913-947 (2003).</li></ol>

著者らは、開示することは何もない。

私たちの手順は、マイクロ流体ベースの電気化学バイオチップの製造およびDNAハイブリダイゼーション事象の分析のためのその利用を実証する。高収量制御微細加工プロセスを通して、私たちは、電気化学変換器のアレイに統合マイクロスケールチャネルから構成デバイスを開発しています。私たちは、反復的なアプローチを通じて、電気化学チップとマイクロ流体チャネル型のフォトリソ?...

マイクロ流体ラボ·オン·チップ（LOC）のデバイスは、臨床診断、環境モニタリングおよび生物医学研究に多くの利点を提供する。これらのデバイスは、処置の多様な試薬の混 ​​合、結合ベースの親和性、シグナル伝達、および細胞培養を含む1-4に行うことができるチップの領域に流体の流れを制御するためのマイクロ流体チャネルを利用する。マイクロフルイディクスは、マイクロウェルプレートリーダーまたは電気泳動ゲルシフトアッセイのような従来の臨床診断ツールに比べて多くの利点を提供します。マイクロ流体デバイスは、同様のアッセイを実行するために大きさの2から3桁（マイクロリットルとは対照的にナノリットル）より少ない試薬を必要とします。また、これらのデバイスは、いくつかの生物学的事象は、チャネル5,6内の種の小さい閉じ込めに起因し発生することにより、速度を増加させることができる。第三に、センサは、無標識de​​tectioを提供することができるリソグラフィ及びエッチング技術を用いて、マイクロ流体デバイス内に統合することができる。nは。最後に、これらのデバイスは製造が安価であり、7〜10を動作させるために、技術者の一部にはほとんどの作業を必要とします。 ラベルフリー検出は、典型的には、光学的または電気的変換器を用いて行われる。光デバイスが原因サンプル中の分析でより低い干渉により良いセンシング性能を提示することができます。それにもかかわらず、その性能は、サンプルのバックグラウンドは、センサ11と同じ共振波長を有する場合において損なわれる。マイクロ流体システムにおいて生物学的および化学的検出を行うために電気信号を使用することには多くの利点がある。これらのセンサは、一般的にのみ作動するようにパターニングされた電極を必要とするため製造が本質的にあまり複雑である。他の信号のモダリティは、信号12-15に変換するための変換器が必要になることがありながら、また、電気信号は、直接、ほとんどの測定機器とインターフェースすることができます。電気センサーは、一般的にimpedancの変化を測定電子16,17、容量18、またはレドックス活性が19。これらのシステムは小型化されるが、新たな課題が提示される。克服すべき最も重要な課題は、（低いサンプル体積及びレイノルズ数に対する）流体の試料調製、混合、（建設資材と分析物との間の毛管力、表面粗さ、化学的相互作用を含む）の物理的及び化学的効果、低い信号 - を20-23、複雑な生物学的サンプル（ 例えば 、血液および唾液）における電気活性分析物からの潜在的な干渉（縮小表面積および体積によって生成される）信号対雑音比。これらの効果の詳しい調査の結果、彼らの全体的なパフォーマンスを改善だろ再現可能な方法で、これらのデバイスの正確な製造および操作のためのガイドラインになります。 DNAハイブリダイゼーションの検出は、遺伝性疾患24,25とCANCのさまざまな形態を診断するために広く使用されているえー26。毎年、インフルエンザの複数の株は、DNAハイブリダイゼーション技術27からの結果を使用している患者において同定される。インフルエンザウイルスは、米国だけで28人が死亡、毎年36,000を占めています。このような例としては、感度または特異性を犠牲にすることなく低いサンプル体積とし、わずかなコストでプレートリーダーまたはゲルシフトアッセイと同じアッセイ技術を実行することができるベンチトップマイクロ流体デバイスから利益を得ることができる。によるラベルフリー電気化学センシングの多くの利点には、DNAハイブリダイゼーション事象29,30の検出のために広く使用されている。 （ミリメートルの範囲内）マクロスケール電極が関心の溶液ビーカー中に浸漬されている設定は、それらの一致する相補的配列への一本鎖DNA配列の結合キネティクスに関する非常に機密性の高いデータを提供するために使用することができる。最近では、MICR電気化学センシングを組み込んでも数少ない進歩があったDNAハイブリダイゼーションのためのofluidics。研究は、ハイブリダイゼーション動力学31とマイクロ流体チャネルにおける検出するためのセンサ15の統合に関して行われてきた。しかし、依然として複雑な試料調製工程なしで並列にDNAハイブリダイゼーション事象を分析することができる迅速なハイスループットマイクロ流体装置の必要性が存在する。 この研究で提示デバイスは、複数の相互作用が並行して、複雑な試料調製工程なしでスクリーニングされるのを可能にするプラットフォームを提供する。私たちのプロトコルは、電気化学バイオチップは、微小電気機械システム（MEMS）技術32,33で微細加工されているかを、マイクロ流体ベースの紹介。私たちは、ポリジメチルシロキサン（PDMS）製のマイクロ流体チップ、および電極のアレイで構成される電気化学チップ、両方の製造プロセスを説明する。一本鎖DNAプローブを用いたバイオチップの化学的機能も対処される。最後に、ABIL具体的なssDNA標的を検出および分析するためのバイオセンサーのITYが実証される。全体的に、マイクロ流体ベースの電気化学バイオチップは、迅速かつハイスループット分析技術である。これは、生物学的分子と導電性の変換器間の相互作用を調査するために使用することができ、ラボオンチップ、さまざまな用途に利用することができる。

<table border="0" cellpadding="0" cellspacing="0" width="765">
	<tbody>
		<tr height="42">
			<td height="42" style="height:42px;width:216px;">Name of Material/ Equipment</td>
			<td style="width:155px;">Company</td>
			<td style="width:227px;">Catalog Number</td>
			<td style="width:167px;">Comments/Description</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">Material</td>
			<td style="width:155px;"></td>
			<td style="width:227px;"></td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">4&#39;&#39; Silicon wafer</td>
			<td style="width:155px;">Ultrasil</td>
			<td style="width:227px;">4-5664</td>
			<td>Single side polished; P-type; Boron doped; Orientation 1-0-0; Thickness 500 micron; Resistance 10-20 ohm*cm</td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:216px;">Shipley 1813 photoresist (&#34;PR1&#34;)</td>
			<td style="width:155px;">Microchem</td>
			<td style="width:227px;"></td>
			<td style="width:167px;">positive photoresist</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">AZ5214 photoresist (&#34;PR2&#34;)</td>
			<td style="width:155px;">Hoechst Celanse</td>
			<td style="width:227px;"></td>
			<td style="width:167px;">positive photoresist</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">SU-8 50 photoresist (&#34;PR3&#34;)</td>
			<td style="width:155px;">Microchem</td>
			<td style="width:227px;"></td>
			<td style="width:167px;">negative photoresist</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">Gold etchant</td>
			<td style="width:155px;">Transene</td>
			<td style="width:227px;">TFA</td>
			<td style="width:167px;">No dilution</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">Chromium etchant</td>
			<td style="width:155px;">Transene</td>
			<td style="width:227px;">1020AC</td>
			<td style="width:167px;">No dilution</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">PDMS elastomer</td>
			<td style="width:155px;">Dow Corning</td>
			<td style="width:227px;">3097366 1004</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">PDMS curing agent</td>
			<td style="width:155px;">Dow Corning</td>
			<td style="width:227px;">3097358 1004</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">Biopsy punching tool</td>
			<td style="width:155px;">Healthlink</td>
			<td style="width:227px;">BP20</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">Tygon flexible tubing</td>
			<td style="width:155px;">Cole Parmer&#160;</td>
			<td style="width:227px;">R 3603</td>
			<td>.015&#34;</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">Tygon flexible adapter</td>
			<td style="width:155px;">Cole Parmer&#160;</td>
			<td style="width:227px;">06417-41</td>
			<td>.0625&#34;</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">1 mL syringe</td>
			<td style="width:155px;">Beckton-Dickenson</td>
			<td align="right" style="width:227px;">301025</td>
			<td></td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:216px;">Monobasic potassium phosphate</td>
			<td style="width:155px;">Fluka</td>
			<td align="right" style="width:227px;">1551139</td>
			<td></td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:216px;">Potassium phosphate dibasic anhydrous</td>
			<td style="width:155px;">Sigma</td>
			<td style="width:227px;">RES20765-A7</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">Sodium chloride</td>
			<td style="width:155px;">Sigma-Aldrich</td>
			<td style="width:227px;">S7653</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">tris(2-carboxyethyl)phosphine</td>
			<td style="width:155px;">Aldrich</td>
			<td style="width:227px;">C4706</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">6-mercapto-1-hexanol</td>
			<td style="width:155px;">Aldrich</td>
			<td align="right" style="width:227px;">451088</td>
			<td></td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:216px;">20x concentrated saline-sodium citrate buffer</td>
			<td style="width:155px;">Sigma</td>
			<td align="right" style="width:227px;">93017</td>
			<td></td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:216px;">Potassium hexacyanoferrate(III)</td>
			<td style="width:155px;">Aldrich</td>
			<td align="right" style="width:227px;">455946</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">Sodium ferrocyanide</td>
			<td style="width:155px;">Aldrich</td>
			<td style="width:227px;">CDS001589</td>
			<td></td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:216px;">Target ssDNA #1</td>
			<td style="width:155px;">Integrated DNA Technologies (IDT)</td>
			<td style="width:227px;">100 nmole DNA oligo</td>
			<td>5&#8217;-/5ThioMC6-D/AAAGCTCCGATAGCGCTCCG 
			TGGACGTCCC-3&#8217;</td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:216px;">Complementary ssDNA #1</td>
			<td style="width:155px;">Integrated DNA Technologies (IDT)</td>
			<td style="width:227px;">100 nmole DNA oligo</td>
			<td>5&#8217;-GGGACGTCCACGGAGCGCTA 
			TCGGAGCTTT-3&#8217;</td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:216px;">Target ssDNA #2</td>
			<td style="width:155px;">Integrated DNA Technologies (IDT)</td>
			<td style="width:227px;">100 nmole DNA oligo</td>
			<td>5&#8217;-/5ThioMC6-D/ACGCGTCAGGTCATTGACGA 
			ATCGATGAGT-3&#8217;</td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:216px;">Complementary ssDNA #2</td>
			<td style="width:155px;">Integrated DNA Technologies (IDT)</td>
			<td style="width:227px;">100 nmole DNA oligo</td>
			<td>5&#8217;-ACTCATCGATTCGTCAATGA 
			CCTGACCCGT-3&#8217;</td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:216px;">Target ssDNA #3</td>
			<td style="width:155px;">Integrated DNA Technologies (IDT)</td>
			<td style="width:227px;">100 nmole DNA oligo</td>
			<td>5&#8217;-/5ThioMC6-D/ACCTAGATCCAGTAGTTAGA 
			CCCATGATGA-3&#8217;</td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:216px;">Complementary ssDNA #3</td>
			<td style="width:155px;">Integrated DNA Technologies (IDT)</td>
			<td style="width:227px;">100 nmole DNA oligo</td>
			<td>5&#8217;-TCATCATGGGTCTAACTACT 
			GGATCTAGGT-3&#8217;</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;"></td>
			<td style="width:155px;"></td>
			<td style="width:227px;"></td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">Instrument</td>
			<td style="width:155px;"></td>
			<td style="width:227px;"></td>
			<td></td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:216px;">Plasma Enhanced Chemical Vapor Deposition (PECVD)</td>
			<td style="width:155px;">Oxford Instruments</td>
			<td style="width:227px;">PlasmaLab System 100</td>
			<td>Chamber pressure 1000 mTorr, Tempreture 200 celsius degrees, RF power 20 W, Gasses: a) N2O flow rate 710 sccm; b) a composition of 5% SiH4 and 95% N2 gasses flow rate 170 sccm, SiO2 growth rate 690 Angstrom/minute.</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">DC sputtering unit</td>
			<td style="width:155px;">AJA International</td>
			<td style="width:227px;">ATC 1800-V</td>
			<td>For Chrome: Chamber pressure 10 mTorr, Argon flow rate 20 sccm, Supplied DC power 200 W, Sputter rate 10 nm/min. For Gold: Chamber pressure 10 mTorr, Argon flow rate 20 sccm, Supplied DC power 200 W, Sputter rate 36 nm/min.</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">E-beam evaporation system</td>
			<td style="width:155px;">Denton</td>
			<td style="width:227px;">Custom-built</td>
			<td>For Titanium: Chamber pressure &#60;2E-6 Torr, Supplied power to the e-gun 7.5k W, Filament current 150-200 mA, Evaporation rate 6-10 Angstrom/second. For Platinum: Chamber pressure &#60;2E-6 Torr, Supplied power to the e-gun 7.5k W, Filament current 150-200 mA, Evaporation rate 2-3 Angstrom/second.&#160;</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">Potentiostat</td>
			<td style="width:155px;">CH Instruments</td>
			<td style="width:227px;">660D</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">Syringe pump</td>
			<td style="width:155px;">KD Scientific</td>
			<td style="width:227px;">KDS230</td>
			<td></td>
		</tr>
	</tbody>
</table>

a microfluidic-based electrochemical biochip for label-free dna hybridization analysis

マイクロスケールに分析ベンチトップ手順の小型化、反応時間、コスト、及び前処理ステップの統合に関して有意な利点を提供する。それはポイント·オブ·ケア各種疾患のバイオマーカーでのリアルタイム評価のための技術を提供するためのDNAハイブリダイゼーション事象の解析に向けて、これらのデバイスを利用することが重要です。しかし、デバイスのフットプリントは、さまざまな物理現象の増加優位性を低下させる。これらの現象は、加工精度や装置の動作信頼性に影響を与える。したがって、正確な全体的なパフォーマンスを改善するために、再現可能な方法でこれらのデバイスを製造し操作する大きな必要性がある。ここでは、プロトコルおよび製造およびDNAハイブリダイゼーション事象の正確な分析のためのマイクロ流体ベースの電気化学バイオチップの動作に用いられる方法を記載する。バイオチップは2つの部分から構成されています。マイクロ流体チップを搭載した3ポリジメチルシロキサン（PDMS）からなる平行マイクロチャネル、及び3×3配列の電気化学的マイクロチップ。 DNAハイブリダイゼーション事象は、電気化学インピーダンス分光法（EIS）分析を用いて検出される。 EISの分析は、これらの長さスケールでの支配的な電気化学システムのプロパティの監視のバリエーションを可能にします。バイオセンサーの両方の電荷移動と拡散抵抗の変化をモニターする能力によって、私達は20分以下、相補鎖DNA標的に対する選択性は3.8 nMでの計算された検出限界、および他の非相補的ssDNAを13％の交差反応性を実証インキュベーション。この方法は、マイクロスケールの領域における拡散の挙動に解明することによって、およびDNAハイブリダイゼーション事象の研究を可能にすることによって、小型化デバイスの性能を向上させることができる。

実験装置のための制御可能かつ正確な製造プロセスは、研究に不可欠である。これは、研究者が再現可能なハイスループット実験を得ることができる。ここでは、マイクロ流体ベースの電気化学バイオチップ（ 図1）の高い収率、高い再現性の微細加工プロセスを実証している。低い故障率で、いくつかのデバイスは、溶液の漏れにつながる接合の問題を示している。バイオチッ...

Watch this Scientific Journal Video about A Microfluidic-based Electrochemical Biochip for Label-free DNA Hybridization Analysis at JoVE.com

A Microfluidic-based Electrochemical Biochip for Label-free DNA Hybridization Analysis

私たちは、DNAハイブリダイゼーション検出のためのマイクロ流体ベースの電気化学バイオチップを提供する。のssDNAプローブ官能化に続いて、特異性、感度、および検出限界は、相補的および非相補的なssDNA標的と研究されている。結果は、3.8 nMでの検出限界で、電気化学システム上のDNAハイブリダイゼーション事象の影響を例示する。

ラベルフリーDNAハイブリダイゼーション分析のためのマイクロ流体ベースの電気化学バイオチップ

Miniaturization of analytical benchtop procedures into the micro-scale provides significant advantages in regards to reaction time, cost, and integration ...

a-microfluidic-based-electrochemical-biochip-for-label-free-dna

Research

JoVE Journal

Bioengineering

17.0K ビュー.  University of Maryland. 私たちは、DNAハイブリダイゼーション検出のためのマイクロ流体ベースの電気化学バイオチップを提供する。のssDNAプローブ官能化に続いて、特異性、感度、および検出限界は、相補的および非相補的なssDNA標的と研究されている。結果は、3.8 nMでの検出限界で、電気化学システム上のDNAハイブリダイゼーション事象の影響を例示する。 

ビデオ: A Microfluidic-based Electrochemical Biochip for Label-free DNA Hybridization Analysis

A Microfluidic-based Hydrodynamic Trap for Single Particles

The ability to confine and manipulate single particles in free solution is a key enabling technology for fundamental and applied science. Methods for particle trapping based on optical, magnetic, electrokinetic, and acoustic techniques have led to major advancements in physics and biology ranging from the molecular to cellular level. In this article, we introduce a new microfluidic-based technique for particle trapping and manipulation based solely on hydrodynamic fluid flow. Using this method, we demonstrate trapping of micro- and nano-scale particles in aqueous solutions for long time scales. The hydrodynamic trap consists of an integrated microfluidic device with a cross-slot channel geometry where two opposing laminar streams converge, thereby generating a planar extensional flow with a fluid stagnation point (zero-velocity point). In this device, particles are confined at the trap center by active control of the flow field to maintain particle position at the fluid stagnation point. In this manner, particles are effectively trapped in free solution using a feedback control algorithm implemented with a custom-built LabVIEW code. The control algorithm consists of image acquisition for a particle in the microfluidic device, followed by particle tracking, determination of particle centroid position, and active adjustment of fluid flow by regulating the pressure applied to an on-chip pneumatic valve using a pressure regulator. In this way, the on-chip dynamic metering valve functions to regulate the relative flow rates in the outlet channels, thereby enabling fine-scale control of stagnation point position and particle trapping. The microfluidic-based hydrodynamic trap exhibits several advantages as a method for particle trapping. Hydrodynamic trapping is possible for any arbitrary particle without specific requirements on the physical or chemical properties of the trapped object. In addition, hydrodynamic trapping enables confinement of a "single" target object in concentrated or crowded particle suspensions, which is difficult using alternative force field-based trapping methods. The hydrodynamic trap is user-friendly, straightforward to implement and may be added to existing microfluidic devices to facilitate trapping and long-time analysis of particles. Overall, the hydrodynamic trap is a new platform for confinement, micromanipulation, and observation of particles without surface immobilization and eliminates the need for potentially perturbative optical, magnetic, and electric fields in the free-solution trapping of small particles.

In this article, we present a microfluidic-based method for particle confinement based on hydrodynamic flow. We demonstrate stable particle trapping at a fluid stagnation point using a feedback control mechanism, thereby enabling confinement and micromanipulation of arbitrary particles in an integrated microdevice.

単一粒子のためのマイクロ流体ベースの流体力学的トラップ

The ability to confine and manipulate single particles in free solution is a key enabling technology for fundamental and applied science.  Methods for ...

生物工学

Fabrication of Electrochemical-DNA Biosensors for the Reagentless Detection of Nucleic Acids, Proteins and Small Molecules

As medicine is currently practiced, doctors send specimens to a central laboratory for testing and thus must wait hours or days to 
receive the results. Many patients would be better served by rapid, bedside tests. To this end our laboratory and others have developed a versatile, reagentless 
biosensor platform that supports the quantitative, reagentless, electrochemical detection of nucleic acids (DNA, RNA), proteins (including antibodies) and small 
molecules analytes directly in unprocessed clinical and environmental samples. In this video, we demonstrate the preparation and use of several biosensors in this 
"E-DNA" class. In particular, we fabricate and demonstrate sensors for the detection of a target DNA sequence in a polymerase chain reaction mixture, an HIV-specific antibody and the drug cocaine. The preparation procedure requires only three hours of hands-on effort followed by an overnight incubation, and their use requires only minutes.

"E-DNA" sensors, reagentless, electrochemical biosensors that perform well even when challenged directly in blood and other complex matrices, have been adapted to the detection of a wide range of nucleic acid, protein and small molecule analytes. Here we present a general procedure for the fabrication and use of such sensors.

核酸、タンパク質と低分子のReagentless検出用電気化学DNAバイオセンサーの作製

As medicine is currently practiced, doctors send specimens to a central laboratory for testing and thus must wait hours or days to 
receive the results. ...

Non-contact, Label-free Monitoring of Cells and Extracellular Matrix using Raman Spectroscopy

Non-destructive, non-contact and label-free technologies to monitor cell and tissue cultures are needed in the field of biomedical research.1-5 However, currently available routine methods require processing steps and alter sample integrity. Raman spectroscopy is a fast method that enables the measurement of biological samples without the need for further processing steps. This laser-based technology detects the inelastic scattering of monochromatic light.6 As every chemical vibration is assigned to a specific Raman band (wavenumber in cm-1), each biological sample features a typical spectral pattern due to their inherent biochemical composition.7-9 Within Raman spectra, the peak intensities correlate with the amount of the present molecular bonds.1 Similarities and differences of the spectral data sets can be detected by employing a multivariate analysis (e.g. principal component analysis (PCA)).10
Here, we perform Raman spectroscopy of living cells and native tissues. Cells are either seeded on glass bottom dishes or kept in suspension under normal cell culture conditions (37 &deg;C, 5% CO2) before measurement. Native tissues are dissected and stored in phosphate buffered saline (PBS) at 4 &deg;C prior measurements. Depending on our experimental set up, we then either focused on the cell nucleus or extracellular matrix (ECM) proteins such as elastin and collagen. For all studies, a minimum of 30 cells or 30 random points of interest within the ECM are measured. Data processing steps included background subtraction and normalization.

Raman spectroscopy is a suitable technique for the non-contact, label-free analysis of living cells, tissue-engineered constructs and native tissues. Source-specific spectral fingerprints can be generated and analyzed using multivariate analysis.

ラマン分光法を用いた細胞と細胞外マトリックスの非接触、ラベルフリーの監視

Non-destructive, non-contact and label-free technologies to monitor cell and tissue cultures are needed in the field of biomedical research.1-5 However, ...

Hollow Microneedle-based Sensor for Multiplexed Transdermal Electrochemical Sensing

The development of a minimally invasive multiplexed monitoring system for rapid analysis of biologically-relevant molecules could offer individuals suffering from chronic medical conditions facile assessment of their immediate physiological state. Furthermore, it could serve as a research tool for analysis of complex, multifactorial medical conditions. In order for such a multianalyte sensor to be realized, it must be minimally invasive, sampling of interstitial fluid must occur without pain or harm to the user, and analysis must be rapid as well as selective.
Initially developed for pain-free drug delivery, microneedles have been used to deliver vaccines and pharmacologic agents (e.g., insulin) through the skin.1-2 Since these devices access the interstitial space, microneedles that are integrated with microelectrodes can be used as transdermal electrochemical sensors. Selective detection of glucose, glutamate, lactate, hydrogen peroxide, and ascorbic acid has been demonstrated using integrated microneedle-electrode devices with carbon fibers, modified carbon pastes, and platinum-coated polymer microneedles serving as transducing elements.3-7,8
This microneedle sensor technology has enabled a novel and sophisticated analytical approach for in situ and simultaneous detection of multiple analytes. Multiplexing offers the possibility of monitoring complex microenvironments, which are otherwise difficult to characterize in a rapid and minimally invasive manner. For example, this technology could be utilized for simultaneous monitoring of extracellular levels of, glucose, lactate and pH,9 which are important metabolic indicators of disease states7,10-14 (e.g., cancer proliferation) and exercise-induced acidosis.15

This article details the construction of a multiplexed microneedle-based sensor. The device is being developed for in situ sampling and electrochemical analysis of multiple analytes in a rapid and selective manner. We envision clinical medicine and biomedical research uses for these microneedle-based sensors.

多重経皮電気化学センシングのための中空マイクロニードルベースの​​センサ

The development of a minimally invasive multiplexed monitoring system for rapid analysis of biologically-relevant molecules could offer individuals ...

Bacterial Detection &amp; Identification Using Electrochemical Sensors

Electrochemical sensors are widely used for rapid and accurate measurement of blood glucose and can be adapted for detection of a wide variety of analytes. Electrochemical sensors operate by transducing a biological recognition event into a useful electrical signal. Signal transduction occurs by coupling the activity of a redox enzyme to an amperometric electrode. Sensor specificity is either an inherent characteristic of the enzyme, glucose oxidase in the case of a glucose sensor, or a product of linkage between the enzyme and an antibody or probe.
Here, we describe an electrochemical sensor assay method to directly detect and identify bacteria. In every case, the probes described here are DNA oligonucleotides. This method is based on sandwich hybridization of capture and detector probes with target ribosomal RNA (rRNA). The capture probe is anchored to the sensor surface, while the detector probe is linked to horseradish peroxidase (HRP). When a substrate such as 3,3',5,5'-tetramethylbenzidine (TMB) is added to an electrode with capture-target-detector complexes bound to its surface, the substrate is oxidized by HRP and reduced by the working electrode. This redox cycle results in shuttling of electrons by the substrate from the electrode to HRP, producing current flow in the electrode.

We describe an electrochemical sensor assay method for rapid bacterial detection and identification. The assay involves a sensor array functionalized with DNA oligonucleotide capture probes for ribosomal RNA (rRNA) species-specific sequences. Sandwich hybridization of target rRNA with the capture probe and a horseradish peroxidase-linked DNA oligonucleotide detector probe produces a measurable amperometric current.

電気化学センサを用いた細菌検出＆識別

Electrochemical  sensors are widely used for rapid and accurate measurement of blood glucose and  can be adapted for detection of a wide variety of ...

Multi-analyte Biochip (MAB) Based on All-solid-state Ion-selective Electrodes (ASSISE) for Physiological Research

Lab-on-a-chip (LOC) applications in environmental, biomedical, agricultural, biological, and spaceflight research require an ion-selective electrode (ISE) that can withstand prolonged storage in complex biological media 1-4. An all-solid-state ion-selective-electrode (ASSISE) is especially attractive for the aforementioned applications. The electrode should have the following favorable characteristics: easy construction, low maintenance, and (potential for) miniaturization, allowing for batch processing. A microfabricated ASSISE intended for quantifying H+, Ca2+, and CO32- ions was constructed. It consists of a noble-metal electrode layer (i.e. Pt), a transduction layer, and an ion-selective membrane (ISM) layer. The transduction layer functions to transduce the concentration-dependent chemical potential of the ion-selective membrane into a measurable electrical signal.
The lifetime of an ASSISE is found to depend on maintaining the potential at the conductive layer/membrane interface 5-7. To extend the ASSISE working lifetime and thereby maintain stable potentials at the interfacial layers, we utilized the conductive polymer (CP) poly(3,4-ethylenedioxythiophene) (PEDOT) 7-9 in place of silver/silver chloride (Ag/AgCl) as the transducer layer. We constructed the ASSISE in a lab-on-a-chip format, which we called the multi-analyte biochip (MAB) (Figure 1).
Calibrations in test solutions demonstrated that the MAB can monitor pH (operational range pH 4-9), CO32- (measured range 0.01 mM - 1 mM), and Ca2+ (log-linear range 0.01 mM to 1 mM). The MAB for pH provides a near-Nernstian slope response after almost one month storage in algal medium. The carbonate biochips show a potentiometric profile similar to that of a conventional ion-selective electrode. Physiological measurements were employed to monitor biological activity of the model system, the microalga Chlorella vulgaris.
The MAB conveys an advantage in size, versatility, and multiplexed analyte sensing capability, making it applicable to many confined monitoring situations, on Earth or in space.
Biochip Design and Experimental Methods
The biochip is 10 x 11 mm in dimension and has 9 ASSISEs designated as working electrodes (WEs) and 5 Ag/AgCl reference electrodes (REs). Each working electrode (WE) is 240 &mu;m in diameter and is equally spaced at 1.4 mm from the REs, which are 480 &mu;m in diameter. These electrodes are connected to electrical contact pads with a dimension of 0.5 mm x 0.5 mm. The schematic is shown in Figure 2.
Cyclic voltammetry (CV) and galvanostatic deposition methods are used to electropolymerize the PEDOT films using a Bioanalytical Systems Inc. (BASI) C3 cell stand (Figure 3). The counter-ion for the PEDOT film is tailored to suit the analyte ion of interest. A PEDOT with poly(styrenesulfonate) counter ion (PEDOT/PSS) is utilized for H+ and CO32-, while one with sulphate (added to the solution as CaSO4) is utilized for Ca2+. The electrochemical properties of the PEDOT-coated WE is analyzed using CVs in redox-active solution (i.e. 2 mM potassium ferricyanide (K3Fe(CN)6)). Based on the CV profile, Randles-Sevcik analysis was used to determine the effective surface area 10. Spin-coating at 1,500 rpm is used to cast ~2 &mu;m thick ion-selective membranes (ISMs) on the MAB working electrodes (WEs).
The MAB is contained in a microfluidic flow-cell chamber filled with a 150 &mu;l volume of algal medium; the contact pads are electrically connected to the BASI system (Figure 4). The photosynthetic activity of Chlorella vulgaris is monitored in ambient light and dark conditions.

Multi-analyte Biochip MAB Based on All-solid-state Ion-selective Electrodes ASSISE for Physiological Research

All-solid-state ion-selective electrodes (ASSISEs) constructed from a conductive polymer (CP) transducer provide several months of functional lifetime in liquid media. Here, we describe the fabrication and calibration process of ASSISEs in a lab-on-a-chip format. The ASSISE is demonstrated to have maintained a near-Nernstian slope profile after prolonged storage in complex biological media.

生理学的研究のための全固体型イオン選択性電極（ASSISE）に基づき、多検体バイオチップ（MAB）

Lab-on-a-chip (LOC) applications in environmental, biomedical, agricultural, biological, and spaceflight research require an ion-selective electrode (ISE) ...

Microfluidic-based Electrotaxis for On-demand Quantitative Analysis of Caenorhabditis elegans' Locomotion

The nematode Caenorhabditis elegans is a versatile model organism for biomedical research because of its conservation of disease-related genes and pathways as well as its ease of cultivation. Several C. elegans disease models have been reported, including neurodegenerative disorders such as Parkinson's disease (PD), which involves the degeneration of dopaminergic (DA) neurons 1. Both transgenes and neurotoxic chemicals have been used to induce DA neurodegeneration and consequent movement defects in worms, allowing for investigations into the basis of neurodegeneration and screens for neuroprotective genes and compounds 2,3.
Screens in lower eukaryotes like C. elegans provide an efficient and economical means to identify compounds and genes affecting neuronal signaling. Conventional screens are typically performed manually and scored by visual inspection; consequently, they are time-consuming and prone to human errors. Additionally, most focus on cellular level analysis while ignoring locomotion, which is an especially important parameter for movement disorders.
We have developed a novel microfluidic screening system (Figure 1) that controls and quantifies C. elegans' locomotion using electric field stimuli inside microchannels. We have shown that a Direct Current (DC) field can robustly induce on-demand locomotion towards the cathode ("electrotaxis") 4. Reversing the field's polarity causes the worm to quickly reverse its direction as well. We have also shown that defects in dopaminergic and other sensory neurons alter the swimming response 5. Therefore, abnormalities in neuronal signaling can be determined using locomotion as a read-out. The movement response can be accurately quantified using a range of parameters such as swimming speed, body bending frequency and reversal time.
Our work has revealed that the electrotactic response varies with age. Specifically, young adults respond to a lower range of electric fields and move faster compared to larvae 4. These findings led us to design a new microfluidic device to passively sort worms by age and phenotype 6.
We have also tested the response of worms to pulsed DC and Alternating Current (AC) electric fields. Pulsed DC fields of various duty cycles effectively generated electrotaxis in both C. elegans and its cousin C. briggsae 7. In another experiment, symmetrical AC fields with frequencies ranging from 1 Hz to 3 KHz immobilized worms inside the channel 8.
Implementation of the electric field in a microfluidic environment enables rapid and automated execution of the electrotaxis assay. This approach promises to facilitate high-throughput genetic and chemical screens for factors affecting neuronal function and viability.

Microfluidic-based Electrotaxis for On-demand Quantitative Analysis of Caenorhabditis elegans&#039; Locomotion

A semi-automated micro-electro-fluidic method to induce on-demand locomotion in Caenorhabditis elegans is described. This method is based on the neurophysiologic phenomenon of worms responding to mild electric fields (&#x201C;electrotaxis&#x201D;) inside microfluidic channels. Microfluidic electrotaxis serves as a rapid, sensitive, low-cost, and scalable technique to screen for factors affecting neuronal health.

のオンデマンド定量分析のためのマイクロ流体ベースの走電性<em&gt;線虫（Caenorhabditis elegans）</em&gt; &#39;ロコモーション

The nematode Caenorhabditis elegans is a versatile model organism for biomedical research because of its conservation of disease-related genes and ...

Use of Label-free Optical Biosensors to Detect Modulation of Potassium Channels by G-protein Coupled Receptors

Ion channels control the electrical properties of neurons and other excitable cell types by selectively allowing ions to flow through the plasma membrane1. To regulate neuronal excitability, the biophysical properties of ion channels are modified by signaling proteins and molecules, which often bind to the channels themselves to form a heteromeric channel complex2,3. Traditional assays examining the interaction between channels and regulatory proteins require exogenous labels that can potentially alter the protein&#39;s behavior and decrease the physiological relevance of the target, while providing little information on the time course of interactions in living cells. Optical biosensors, such as the X-BODY Biosciences BIND Scanner system, use a novel label-free technology, resonance wavelength grating (RWG) optical biosensors, to detect changes in resonant reflected light near the biosensor. This assay allows the detection of the relative change in mass within the bottom portion of living cells adherent to the biosensor surface resulting from ligand induced changes in cell adhesion and spreading, toxicity, proliferation, and changes in protein-protein interactions near the plasma membrane. RWG optical biosensors have been used to detect changes in mass near the plasma membrane of cells following activation of G protein-coupled receptors (GPCRs), receptor tyrosine kinases, and other cell surface receptors. Ligand-induced changes in ion channel-protein interactions can also be studied using this assay. In this paper, we will describe the experimental procedure used to detect the modulation of Slack-B sodium-activated potassium (KNa) channels by GPCRs.

Optical biosensor techniques can detect changes in mass near the plasma membrane in living cells and allow one to follow cellular responses in both individual cells and populations of cells. This protocol will describe detection of the modulation of potassium channels by G-protein coupled receptors in intact cells using this approach.

Gタンパク質共役受容体によるカリウムチャネルの調節を検出するために、ラベルフリー光学バイオセンサの使用

Ion channels control the electrical properties of neurons and other excitable cell types by selectively allowing ions to flow through the plasma ...

A Label-free Technique for the Spatio-temporal Imaging of Single Cell Secretions

Inter-cellular communication is an integral part of a complex system that helps in maintaining basic cellular activities. As a result, the malfunctioning of such signaling can lead to many disorders. To understand cell-to-cell signaling, it is essential to study the spatial and temporal nature of the secreted molecules from the cell without disturbing the local environment. Various assays have been developed to study protein secretion, however, these methods are typically based on fluorescent probes which disrupt the relevant signaling pathways. To overcome this limitation, a label-free technique is required.
In this paper, we describe the fabrication and application of a label-free localized surface plasmon resonance imaging (LSPRi) technology capable of detecting protein secretions from a single cell. The plasmonic nanostructures are lithographically patterned onto a standard glass coverslip and can be excited using visible light on commercially available light microscopes. Only a small fraction of the coverslip is covered by the nanostructures and hence this technique is well suited for combining common techniques such as fluorescence and bright-field imaging.
A multidisciplinary approach is used in this protocol which incorporates sensor nanofabrication and subsequent biofunctionalization, binding kinetics characterization of ligand and analyte, the integration of the chip and live cells, and the analysis of the measured signal. As a whole, this technology enables a general label-free approach towards mapping cellular secretions and correlating them with the responses of nearby cells.

Inter-cellular communication is critical for controlling various physiological activities within and outside the cell. This paper describes a protocol for measuring the spatio-temporal nature of single cell secretions. To achieve this, a multidisciplinary approach is used which integrates label-free nanoplasmonic sensing with live cell imaging.

単一細胞分泌物の時空間イメージング用ラベルフリー技術

Inter-cellular communication is an integral part of a complex system that helps in maintaining basic cellular activities. As a result, the malfunctioning ...

Label-free Single Molecule Detection Using Microtoroid Optical Resonators

Detecting small concentrations of molecules down to the single molecule limit has impact on areas such as early detection of disease, and fundamental studies on the behavior of molecules. Single molecule detection techniques commonly utilize labels such as fluorescent tags or quantum dots, however, labels are not always available, increase cost and complexity, and can perturb the events being studied. Optical resonators have emerged as a promising means to detect single molecules without the use of labels. Currently the smallest particle detected by a non-plasmonically-enhanced bare optical resonator system in solution is a 25 nm polystyrene sphere1. We have developed a technique known as Frequency Locking Optical Whispering Evanescent Resonator (FLOWER) that can surpass this limit and achieve label-free single molecule detection in aqueous solution2. As signal strength scales with particle volume, our work represents a &#62; 100x improvement in the signal to noise ratio (SNR) over the current state of the art. Here the procedures behind FLOWER are presented in an effort to increase its usage in the field.

We have developed a label-free biosensing system based on optical resonator technology known as Frequency Locking Optical Whispering Evanescent Resonator (FLOWER) that is capable of detecting single molecules in solution. Here the procedures behind this work are described and presented.

マイクロトロイドの光共振器を用いたラベルフリー単一分子検出

Detecting small concentrations of molecules down to the single molecule limit has impact on areas such as early detection of disease, and fundamental ...