Name: dynamic electrochemical measurement of chloride ions
Uploaded: 2016-02-05T13:00:00.000+00:00
Duration: 7 min 32 s
Description: Watch this Scientific Journal Video about Dynamic Electrochemical Measurement of Chloride Ions at JoVE.com

This work is a part of the STW project "Integral solution for sustainable construction (IS2C, Fleur van Rossem for her support during the chip fabrication, Justyna Wiedemair for the chip design and Allison Bidulock for her support during the manuscript preparation.

1.チップ製造注：チップは、Ag / AgClの作用電極（WE）、銀/塩化銀擬似参照電極（擬似RE）とガラスチップ上に白金対向電極から構成されています。銀金属16を処理し 、標準的なクリーンルームを使用して、ガラスチップ上に堆積されます。次いで、それを表面上に塩化銀層を形成するために30秒間の0.1MのFeCl 3溶液中に塩素化されています。銀/塩化銀WE（面積= 9.812 平方ミリメートル）は 、図1に示すように、銀/塩化銀疑似REに囲まれ、中央部に位置しています。 <ol><li>平面銀電極を形成するために、ガラス上に金属銀を析出。次のようにクリーンルームでのステップバイステップのチップ製造を与えられます： <ol><li>リソグラフィー<ol><li>ウェハ洗浄のために、濃に最初のガラスウェハを浸します。表面をきれいにするために任意の有機汚染物を除去した後、1 M KOHにするHNO 3風呂。 </li><li> （厚さ200nm）にポジ型レジスト写真を適用しますスピンコーティングによりウエハ、15分間95℃でレジストをプリベーク。 </li><li>ガラスマスクを介して、5秒間UV光にウェハを公開します。 </li><li>現像液中​​のフォトレジストを開発します。これは、電極が堆積され、残りの領域をレジスト未露光によってカバーされるガラスの面積を公開します。 </li></ol></li><li>ガラスエッチング<ol><li>ガラスの露出領域をエッチングするためにKOHの浴に開発ガラスウェーハを浸します。 </li></ol></li><li>金属蒸着（スパッタリング） <ol><li>露出したガラスエリアでの預金銀金属。 <ol><li>第一のガラスのための接着層として20nmのチタン（スパッタリングによって）チタン（Ti）層を堆積させます。 </li><li>次いで、銀とTiとの間の拡散バリアとして（スパッタリングによって）70n​​mのパラジウム（Pd）層を堆積させます。 </li><li>最後に、銀金属の500 nmのを堆積させます。これは、堆積された金属の厚さです。作用電極の面積を4mm 2です。 </li><li> Rを削除しますアセトン浴中でリフトオフ法によりレジストをemaining。 </li></ol></li><li>シングルチップサイズにウェハをDICE。チップは現在、オンチップのAg / AgCl電極を形成するために塩素化される準備ができています。 </li></ol></li></ol></li><li> 図1に示すように、社内で設計されたポリテトラフルオロエチレンチップホルダにチップを置きます。ホルダーは、プッシュピン（またはバネ式のピン）を介して容易に電気的接続を提供し、電気化学セルが含まれています。 </li><li>チップ上のAg / AgCl電極を形成する銀電極を塩化物にします。そのために、30秒間、細胞中の0.1MのFeCl 3溶液を注ぎます。または電極のシルバー色まで暗い灰色がかったなり。短い浸漬時間は、Ag / AgClでの析出につながることができ、一方、より長い浸漬は重要ではありません。 </li><li>残りのFeCl 3【選択チップをきれいにするために脱イオン水でチップをすすぐ今クロノ測定に使用する準備が整いました。 </li></ol>2.電解準備注：電解質は、0.5M KNO 3における塩化カリウムの異なる濃度で調製しました。 <ol><li> 50のKCl電解質原液（溶液50ml中のKClの0.186グラム）の50ミリリットルを準備します。 </li><li>同じ溶液中0.5 M KNO 3バックグラウンド電解質を形成するために、KNO 3の2.52グラムを追加します。 </li><li>脱イオン水50mlにKNO 3の2.52グラムを追加し、他の言葉ではM. 0.5にゼロとKNO 3に塩化物の出発濃度を保管してください。 </li><li>以下の手順に示すように、体系的に、電解質（1ミリモルから6ミリメートル）の塩化物イオン濃度を増加させるための50mMのKClを加えます。 <ol><li> 1 mMの[CL]を取得するために、0.5M KNO 3の4.9ミリリットルに50mMのKClを100μlのを追加します。 </li><li> 2 mMの[CL]を取得するステップ2.4​​.1の溶液に50mMのKCl溶液の104μLを加えます。 </li><li> sの中に50mMのKCl溶液の108.41μLを追加3 mMの[CL]を取得するためのステップ2.4​​.2のolution。 </li><li> 4 mMの[CL]を取得するステップ2.4​​.3の溶液に50mMのKCl溶液の113.19μLを加えます。 </li><li> 5 mMの[CL]を取得するステップ2.4​​.4の溶液に50mMのKCl溶液の118.2μLを加えます。 </li><li> 6 mMの[CL]を取得するステップ2.4​​.5の溶液に50mMのKCl溶液の123.3μLを加えます。 </li></ol></li></ol> 3.実験のセットアップ注：測定は、ポテンショスタット（生物学的科学器具、フランス）を用いて行きました。 <ol><li>チップホルダにチップを置き、ポテンショスタットの対応する端子に接続します。 </li><li> Ag / AgCl電極をポテンショスタットの作用電極端子を接続します。 </li><li>別のAg / AgCl電極をポテンショスタットの参照電極端子を接続します。 </li><li>白金電極にポテンショスタットの対向電極端子を接続します。 </li><li>カンプを配置周囲の騒音を避けるために、ファラデーケージ内LETE実験のセットアップ。 </li></ol> 4.機器の操作設定注：電気化学セルの動作パラメータは、ポテンシオスタットのユーザインタフェースを介して制御されます。印加電流パルス中のAg / AgClをWEの電位応答は、塩化物イオンの異なる濃度について測定されます。 <ol><li>ポテンショスタットのプログラムでは、技術のクロノを開き、動作パラメータを設定します。 <ol><li>要件に応じて次のパラメータを設定します。 <ol><li>電流（I S） </li><li>印加電流（T 秒 ）の時間</li></ol></li><li>ポテンショスタットプログラムで再生ボタンをクリックすることで、この技術を起動します。 注：ポテンショスタットプログラムが適用される電流パルスの下で時間の関数としての潜在的な応答が表示されます。 </li></ol></li></ol> 5.測定とデータAnalysiの<ol><li>電解液中のClイオン濃度対遷移時間の検量線を描画します。 <ol><li>系統的に、塩化カリウム電解液中のCl濃度を変更します。 <ol><li>電気化学セル中の0.5M KNO 3バックグラウンド電解質と1のKClの5ミリリットルで測定を開始します。 </li><li>ポテンショスタットを使用してクロノ実験を開始し、10ミリアンペア/ cm -2で10秒間の電流を印加し、データを格納します。 </li><li>系統的に1 mMの刻みで6 mmの濃度を変更し、測定を繰り返します。 </li></ol></li><li> a.mptファイルとして測定されたデータを格納し、データ処理プログラムのデータファイルを分析します。 <ol><li>電位応答の一次導関数を計算し、一次微分のピークを計算するために社内開発されたデータ処理プログラムを使用してデータを分析します。一次導関数のピークは、遷移時間です。 注：完全なタラをデータ処理プログラムのINGはのアッバスら、2014補足情報に記載されています。 </li></ol></li><li> 1時間ごとの間隔で測定を3回繰り返します。 </li><li>データ処理プログラムですべてのデータファイルを開き、各測定のための遷移時間を計算します。遷移時間計算の詳細手順は以下のとおりであります： <ol><li>クロノ計測のデータファイルから取得した対時間の電位差をプロットします。 </li><li>電位応答の一次導関数を計算します。 </li><li>一次導関数の最大値と、それの時間を示します。一次導関数の最大値の時間は、遷移時間です。 </li></ol></li><li>検量線は、CLイオン濃度に対する遷移時間の平方根をプロット。 </li><li>測定したデータに沿って砂の式25に基づく理論曲線をプロットします。拡散coefficを逆算データプロットからient。 <ol><li>遷移時間との間で測定されたデータプロットで[CL]ライン上の任意の点を取ると、遷移時間の値と[CL]を記録します。砂の方程式は以下のように与えられます。 τ=（FC * / 2J（1-T のCl - ））2Dπ トランスポート番号であり、Dは拡散係数である-ここで、C *は、バルクのClイオン濃度であり、Fはファラデー定数、Jは電流密度、T Clです 。ネグレクトトンのCl -それは、より高いバックグラウンド電解質濃度のためにゼロに近づくので。 </li><li>拡散係数の砂方程式で唯一残っている変数。遷移時間、塩化物イオン濃度および拡散係数の値を取得砂式の印加電流値を置きます。 </li></ol></li></ol></li><li>ドリフト測定<ol><li>中のKClの1 mMと5ミリリットルを注ぎます電気化学セル。 </li><li>ポテンショスタットでは、現在適用されているが、午前10時-2、時間は10秒となるように設定します。 </li><li>測定間の3時間の間隔で3つの測定毎日で2週間のための潜在的な応答を測定します。 </li><li>電解質、 すなわち、1のKCl、測定を行う前に、毎日更新します。 </li><li>測定値の2週間にわたって遷移時間をプロットします。一日あたりの遷移時間の変化は、遷移時間応答のドリフトです。 </li></ol></li><li>遷移時間測定の擬似参照電極の影響 注：このような銀/塩化銀線、白金線と鋼線のような様々な擬似参照は、遷移時間を測定するために試験されます。 <ol><li>電気化学セルでのKCl電解質の4 mMのを注ぎます。 </li><li>擬似参照電極として銀/塩化銀を使用し、ポテンシオスタットの参照電極端子に接続します。陽極電流パルスのCl＆＃へ713;イオンは、Ag / AgClの作用電極の近くに空乏化されます。 </li><li> 10秒間15アム-2の電流密度を印加することにより、クロノ測定を行います。 </li><li>擬似参照電極として白金で測定を繰り返し、データを記録します。 </li><li>擬似参照電極としての鋼棒を用いた測定を繰り返し、データを記録します。 </li><li>使用される様々な擬似参照電極のための測定された遷移時間をプロットします。 </li><li> KCl 5mMのと測定の完全なセットを繰り返し、様々な擬似参照電極を使用して遷移時間をプロットします。 </li></ol></li></ol>

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The authors have nothing to disclose.

遷移時間は変曲の瞬間です。それは、基準電位、すなわち、参照電極の理論的に独立しています。したがって、任意の金属線は、遷移時間測定のための擬似参照電極として用いることができます。コンクリート中の塩化物イオンの既存の電位差測定とは対照的に、この方法は、長期的かつキャリブレーションのない測定を可能にします。さらに感度及び濃度の検出範囲は、印加される?...

Ag / AgCl電極の遷移時間測定に基づいて、塩化物イオンセンサが提供されます。目的は、電解液中の塩化物イオンの長時間連続監視中に固有のドリフトを回避することです。 Ag / AgCl電極の動的な測定手法であるクロノポテンシオメトリー測定は、この目的のために使用されます。ここでのAg / AgCl電極の電位の変化の割合は、刺激（定電流パルス）の間に測定されます。このアプローチの利点は、次のような現場アプリケーション、従って長期（数年）のためのClイオン濃度の検出を可能にする、液体接合参照電極を逃亡し、代わりに、擬似参照電極としての金属製のワイヤを使用し、 で実証されていますコンクリート構造物内部の測定。 コンクリート構造物中の塩化物イオンが劣化1,2の主要な原因の1つです。これは、補強鋼aで孔食開始します構造3の最終的な失敗でND結果。そのため、コンクリート中のClイオンを測定することは、構造4,5の寿命やメンテナンスサイクルを予測することは避けられません。異なる感知原理は、電気化学的な6,7、光学8,9電磁10,11として、コンクリート中の塩化物イオンの測定のために報告されています。しかし、かさばるセットアップを持っている光学、電磁方法、スタンドアロンシステムとして統合し、選択性12の問題を持ってすることは困難です。電気化学的手法では、銀/塩化銀電極の電位差測定は、技術のアプローチ6,7,13の状態です。有望な結果にもかかわらず、このアプローチは、欠陥のあるデータ14,15における基準電位と拡散電位降下結果のドリフトので、実験室規模の測定に限定されています。動的電気化学測定（DEM）に基づいて、遷移時間のアプローチは、潜在的に問題を軽減することができ16ドリフト 。 DEMでは、適用される刺激に対するシステムの応答は、17から19を測定しました。そのようなシステムの例は、クロノポテンショメトリーです。ここで適用される電流パルスは、電極表面近くにイオンを枯渇させる刺激として使用され、対応する電位応答が測定されます。 Ag / AgCl電極で陽極電流がファラデー反応を開始銀（Ag + CL <img alt="図1" src="/files/ftp_upload/53312/arrow.jpg" /> AgCl + e）は、電極表面近くのClイオンの枯渇をもたらします。電位変化は、印加される電流の関数であり、電解質12,20で（選択）イオンの濃度です。現時点では、これらのイオンは、変曲点21を与え 、完全に電極表面近傍に急速に電位上昇の変化率を枯渇させます。電位 - 時間応答曲線（クロノポテンショグラム）上の変曲点は、​​遷移時間を示すから決定することができます電位応答22の一次導関数の最大値。遷移時間は、イオン濃度の特徴です。このアプローチは、異なるイオン濃度17および電解質23,24のpHを決定するために使用されています。 （電流が印加される）作用電極としてのAg / AgCl電極の場合に枯渇イオンは、塩化物イオン17あろう。したがって、その遷移時間を測定し、その濃度を決定します。

<table><tbody><tr><td>Platinum wire (&amp;ge;99.99% trace metals)</td><td>Sigma Aldrich, the Netherlands</td><td>EP1330-1EA</td><td></td></tr><tr><td>Potassium chloride (BioXtra, &amp;ge;99.0%)</td><td>Sigma Aldrich, the Netherlands</td><td>P9333-500G</td><td></td></tr><tr><td>Potassium hydroxide (90% pure reagent grade)</td><td>Sigma Aldrich, the Netherlands</td><td>484016-1KG</td><td></td></tr><tr><td>Ferric chloride&amp;nbsp;</td><td>Sigma Aldrich, the Netherlands</td><td>451649-1G</td><td></td></tr><tr><td>potassium nitrate (&gt;99% reagent grade)&amp;nbsp;</td><td>Sigma Aldrich, the Netherlands</td><td>P6083-500G</td><td></td></tr><tr><td>Ag/AgCl liquid junction reference electrode&amp;nbsp;</td><td>BASi, USA</td><td>model MF-2079</td><td></td></tr><tr><td>VSP potentiostat</td><td>Biologic Science Instruments, France</td><td>VSP 300</td><td></td></tr><tr><td>Steel wire</td><td>Microlab TU Delft</td><td></td><td></td></tr><tr><td>Silver wire&amp;nbsp;</td><td>Sigma Aldrich, the Netherlands</td><td></td><td></td></tr></tbody></table>

dynamic electrochemical measurement of chloride ions

This protocol describes the dynamic measurement of chloride ions using the transition time of a silver silver chloride (Ag/AgCl) electrode. Silver silver chloride electrode is used extensively for potentiometric measurement of chloride ions concentration in electrolyte. In this measurement, long-term and continuous monitoring is limited due to the inherent drift and the requirement of a stable reference electrode. We utilized the chronopotentiometric approach to minimize drift and avoid the use of a conventional reference electrode. A galvanostatic pulse is applied to an Ag/AgCl electrode which initiates a faradic reaction depleting the Cl&#713; ions near the electrode surface. The transition time, which is the time to completely deplete the ions near the electrode surface, is a function of the ion concentration, given by the Nernst equation. The square root of the transition time is in linear relation to the chloride ion concentration. Drift of the response over two weeks is negligible (59 &#181;M/day) when measuring 1 mM [Cl&#713;]using a current pulse of 10 Am-2. This is a dynamic measurement where the moment of transition time determines the response and thus is independent of the absolute potential. Any metal wire can be used as a pseudo-reference electrode, making this approach feasible for long-term measurement inside concrete structures.

Ag / AgCl電極は、標準的なクリーンルームのプロセスを用いてガラスチップ（ 図1.）上に製造されます。クロノポテンシオメトリー測定セットアップ（ 図2）を使用し、応答をポテンシオスタットを用いて測定しました。遷移時間でのClイオン濃度の影響を観察するために、0.5 M KNO 3バックグラウンド中のClイオンの4、5及び6ミリモルを?...

Watch this Scientific Journal Video about Dynamic Electrochemical Measurement of Chloride Ions at JoVE.com

Dynamic Electrochemical Measurement of Chloride Ions

Dynamic measurement of chloride ions is presented. Transition time of an Ag/AgCl electrode, during a chronopotentiometric technique, can give the concentration of chloride ions in electrolyte. This method does not require a stable conventional reference electrode.

塩化物イオンの動的電気化学測定

This protocol describes the dynamic measurement of chloride ions using the transition time of a silver silver chloride (Ag/AgCl) electrode. Silver silver ...

dynamic-electrochemical-measurement-of-chloride-ions

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Electrochemical Analyses and Redox Titration

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11.4K ビュー.  University of Twente. Dynamic measurement of chloride ions is presented. Transition time of an Ag/AgCl electrode, during a chronopotentiometric technique, can give the concentration of chloride ions in electrolyte. This method does not require a stable conventional reference electrode.

ビデオ: Dynamic Electrochemical Measurement of Chloride Ions

Measurement of Ion Concentration in the Unstirred Boundary Layer with Open Patch-Clamp Pipette: Implications in Control of Ion Channels by Fluid Flow

Fluid flow is an important environmental stimulus that controls many physiological and pathological processes, such as fluid flow-induced vasodilation. Although the molecular mechanisms for the biological responses to fluid flow/shear force are not fully understood, fluid flow-mediated regulation of ion channel gating may contribute critically. Therefore, fluid flow/shear force sensitivity of ion channels has been studied using the patch-clamp technique. However, depending on the experimental protocol, the outcomes and interpretation of data can be erroneous. Here, we present experimental and theoretical evidence for fluid flow-related errors and provide methods for estimating, preventing, and correcting these errors. Changes in junction potential between the Ag/AgCl reference electrode and bathing fluid were measured with an open pipette filled with 3 M KCl. Fluid flow could then shift the liquid/metal junction potential to approximately 7 mV. Conversely, by measuring the voltage shift induced by fluid flow, we estimated the ion concentration in the unstirred boundary layer. In the static condition, the real ion concentrations adjacent to the Ag/AgCl reference electrode or ion channel inlet at the cell-membrane surface can reach as low as approximately 30% of that in the flow condition. Placing an agarose 3 M KCl bridge between the bathing fluid and reference electrode may have prevented this problem of junction potential shifting. However, the unstirred layer effect adjacent to the cell membrane surface could not be fixed in this way. Here, we provide a method for measuring real ion concentrations in the unstirred boundary layer with an open patch-clamp pipette, emphasizing the importance of using an agarose salt-bridge while studying fluid flow-induced regulation of ion currents. Therefore, this novel approach, which takes into consideration the real concentrations of ions in the unstirred boundary layer, may provide useful insight on the experimental design and data interpretation related to fluid shear stress regulation of ion channels.

Mechanosensitive ion channels are often studied in terms of fluid flow/shear force sensitivity with patch-clamp recording. However, depending on the experimental protocol, the outcome on fluid flow-regulations of ion channels can be erroneous. Here, we provide methods for preventing and correcting such errors with a theoretical basis.

オープン パッチ ・ クランプのピペットで非撹拌の境界層におけるイオン濃度の測定: 流体によるイオン チャネルの制御の流れ

Fluid flow is an important environmental stimulus that controls many physiological and pathological processes, such as fluid flow-induced vasodilation. ...

生化学

Experimental Protocol to Determine the Chloride Threshold Value for Corrosion in Samples Taken from Reinforced Concrete Structures

The aging of reinforced concrete infrastructure in developed countries imposes an urgent need for methods to reliably assess the condition of these structures. Corrosion of the embedded reinforcing steel is the most frequent cause for degradation. While it is well known that the ability of a structure to withstand corrosion depends strongly on factors such as the materials used or the age, it is common practice to rely on threshold values stipulated in standards or textbooks. These threshold values for corrosion initiation (Ccrit) are independent of the actual properties of a certain structure, which clearly limits the accuracy of condition assessments and service life predictions. The practice of using tabulated values can be traced to the lack of reliable methods to determine Ccrit on-site and in the laboratory.
Here, an experimental protocol to determine Ccrit for individual engineering structures or structural members is presented. A number of reinforced concrete samples are taken from structures and laboratory corrosion testing is performed. The main advantage of this method is that it ensures real conditions concerning parameters that are well known to greatly influence Ccrit, such as the steel-concrete interface, which cannot be representatively mimicked in laboratory-produced samples. At the same time, the accelerated corrosion test in the laboratory permits the reliable determination of Ccrit prior to corrosion initiation on the tested structure; this is a major advantage over all common condition assessment methods that only permit estimating the conditions for corrosion after initiation, i.e., when the structure is already damaged.
The protocol yields the statistical distribution of Ccrit for the tested structure. This serves as a basis for probabilistic prediction models for the remaining time to corrosion, which is needed for maintenance planning. This method can potentially be used in material testing of civil infrastructures, similar to established methods used for mechanical testing.

We propose a method to measure a parameter that is highly relevant for corrosion assessments or predictions of reinforced concrete structures, with the main advantage of permitting testing of samples from engineering structures. This ensures real conditions at the steel-concrete interface, which are crucial to avoid artifacts of laboratory-made samples.

鉄筋コンクリート構造物から採取された検体で腐食の塩化しきい値値を決定する実験的プロトコル

The aging of reinforced concrete infrastructure in developed countries imposes an urgent need for methods to reliably assess the condition of these ...

工学

Electrochemical Roughening of Thin-Film Platinum Macro and Microelectrodes

This protocol demonstrates a method for electrochemical roughening of thin-film platinum electrodes without preferential dissolution at grain boundaries of the metal. Using this method, a crack free, thin-film macroelectrode surface with up to 40 times increase in active surface area was obtained. The roughening is easy to do in a standard electrochemical characterization laboratory and incudes the application of voltage pulses followed by extended application of a reductive voltage in a perchloric acid solution. The protocol includes the chemical and electrochemical preparation of both a macroscale (1.2 mm diameter) and microscale (20 &#181;m diameter) platinum disc electrode surface, roughening the electrode surface and characterizing the effects of surface roughening on electrode active surface area. This electrochemical characterization includes cyclic voltammetry and impedance spectroscopy and is demonstrated for both the macroelectrodes and the microelectrodes. Roughening increases electrode active surface area, decreases electrode impedance, increases platinum charge injection limits to those of titanium nitride electrodes of same geometry and improves substrates for adhesion of electrochemically deposited films.

This protocol demonstrates a method for electrochemical roughening of thin-film platinum electrodes without preferential dissolution at grain boundaries. The electrochemical techniques of cyclic voltammetry and impedance spectroscopy are demonstrated to characterize these electrode surfaces.

薄膜白金マクロ及びマイクロ電極の電気化学粗化

This protocol demonstrates a method for electrochemical roughening of thin-film platinum electrodes without preferential dissolution at grain boundaries ...

化学

Merging Ion Concentration Polarization between Juxtaposed Ion Exchange Membranes to Block the Propagation of the Polarization Zone

The ion concentration polarization (ICP) phenomenon is one of the most prevailing methods to preconcentrate low-abundance biological samples. The ICP induces a noninvasive region for charged biomolecules (i.e., the ion depletion zone), and targets can be preconcentrated on this region boundary. Despite the high preconcentration performances with ICP, it is difficult to find the operating conditions of non-propagating ion depletion zones. To overcome this narrow operating window, we recently developed a new platform for spatiotemporally fixed preconcentration. Unlike preceding methods that only use ion depletion, this platform also uses the opposite polarity of the ICP (i.e., ion enrichment) to stop the propagation of the ion depletion zone. By confronting the enrichment zone with the depletion zone, the two zones merge together and stop. In this paper, we describe a detailed experimental protocol to build this spatiotemporally defined ICP platform and characterize the preconcentration dynamics of the new platform by comparing them with those of the conventional device. Qualitative ion concentration profiles and current-time responses successfully capture the different dynamics between the merged ICP and the stand-alone ICP. In contrast to the conventional one that can fix the preconcentration location at only ~5 V, the new platform can produce a target-condensed plug at a specific location in the broad ranges of operating conditions: voltage (0.5-100 V), ionic strength (1-100 mM), and pH (3.7-10.3).

The protocol for a novel ion concentration polarization (ICP) platform that can stop the propagation of the ICP zone, regardless of the operating conditions is described. This unique ability of the platform lies in the use of merging ion depletion and enrichment, which are two polarities of the ICP phenomenon.

偏光ゾーンの伝播を遮断するように並置されたイオン交換膜との間のイオン濃度分極のマージ

The ion concentration polarization (ICP) phenomenon is one of the most prevailing methods to preconcentrate low-abundance biological samples. The ICP ...

生物工学

Precise Electrochemical Sizing of Individual Electro-Inactive Particles

Nanoimpact electrochemistry enables the time-resolved in situ characterization (e.g., size, catalytic activity) of single nanomaterial units, providing a means of elucidating heterogeneities that would be masked in ensemble studies. To implement this technique with redox inactive particles, a solution-phase redox reaction is used to produce a steady-state background current on a disk ultramicroelectrode. When a particle adsorbs onto the electrode, it produces a stepwise reduction in the exposed electrode area, which produces, in turn, a stepwise decrease in the current commensurate with the size of the adsorbing species. Historically, however, nanoimpact electrochemistry has suffered from &#34;edge effects,&#34; in which the radial diffusion layer formed at the circumference of the ultramicroelectrodes renders the step size dependent not only on the size of the particle but also on where it lands on the electrode. The introduction of electrocatalytic current generation, however, mitigates the heterogeneity caused by edge effects, thus improving the measurement precision. In this approach, termed &#34;electrocatalytic interruption,&#34; a substrate that regenerates the redox probe at the diffusion layer is introduced. This shifts the rate-limiting step of the current generation from diffusion to the homogeneous reaction rate constant, thus reducing flux heterogeneity and increasing the precision of particle sizing by an order of magnitude. The protocol described here explains the set-up and data collection employed in nanoimpact experiments implementing this effect for improved precision in the sizing of redox in-active materials.

As an analytical technique, nanoimpact electrochemistry, an increasingly important approach to counting and characterizing nanometer-scale, electro-inactive particles, suffers from poor precision due to the heterogeneous current distributions that arise from its use of ultramicroelectrodes. Outlined here is a generalized approach, termed "electrocatalytic interruption," that enhances precision in such measurements.

個々の電気不活性粒子の正確な電気化学的サイジング

Nanoimpact electrochemistry enables the time-resolved in situ characterization (e.g., size, catalytic activity) of single nanomaterial units, providing a ...

Double-barreled and Concentric Microelectrodes for Measurement of Extracellular Ion Signals in Brain Tissue

Electrical activity in the brain is accompanied by significant ion fluxes across membranes, resulting in complex changes in the extracellular concentration of all major ions. As these ion shifts bear significant functional consequences, their quantitative determination is often required to understand the function and dysfunction of neural networks under physiological and pathophysiological conditions. In the present study, we demonstrate the fabrication and calibration of double-barreled ion-selective microelectrodes, which have proven to be excellent tools for such measurements in brain tissue. Moreover, so-called &#8220;concentric&#8221; ion-selective microelectrodes are also described, which, based on their different design, offer a far better temporal resolution of fast ion changes. We then show how these electrodes can be employed in acute brain slice preparations of the mouse hippocampus. Using double-barreled, potassium-selective microelectrodes, changes in the extracellular potassium concentration ([K+]o) in response to exogenous application of glutamate receptor agonists or during epileptiform activity are demonstrated. Furthermore, we illustrate the response characteristics of sodium-sensitive, double-barreled and concentric electrodes and compare their detection of changes in the extracellular sodium concentration ([Na+]o) evoked by bath or pressure application of drugs. These measurements show that while response amplitudes are similar, the concentric sodium microelectrodes display a superior signal-to-noise ratio and response time as compared to the double-barreled design. Generally, the demonstrated procedures will be easily transferable to measurement of other ions species, including pH or calcium, and will also be applicable to other preparations.

We demonstrate the fabrication, calibration and properties of two types of ion-selective microelectrodes (double-barreled and concentric) for measurement of ion concentrations in brain tissue. These are then used in the mouse hippocampal slice preparation to show that excitatory activity changes both extracellular potassium and sodium concentrations.

脳組織における測定細胞外イオンの信号のための、二連と同心微小電極

Electrical activity in the brain is accompanied by significant ion fluxes across membranes, resulting in complex changes in the extracellular ...

神経科学

A Micro-agar Salt Bridge Electrode for Analyzing the Proton Turnover Rate of Recombinant Membrane Proteins

To date, more than 50% of all pharmacological drugs target the transport kinetics of membrane proteins. The electrophysiological characterization of membrane carrier proteins reconstituted in lipid bilayer membranes is a powerful but delicate method for the assessment of their physicochemical and pharmacological properties. The substrate turnover number is a unique parameter that allows the comparison of the activity of different membrane proteins. In an electrogenic transport, the gradient of the translocated substrate creates a membrane potential that directly correlates to the substrate turnover rate of the protein. By using silver chloride electrodes, a diffusion potential, also called liquid junction potential, is induced, which alters electrode potential and significantly disturbs precise membrane potential measurements. Diffusion potential can be minimized by a salt bridge, which balances electrode potential. In this article, a micro-agar salt bridge is designed to improve the electrophysiological set-up, which uses micropipettes for the membrane formation. The salt solution is filled into a microcapillary pipette tip, stabilized by the addition of agarose, and can be easily mounted to a standard electrode. The electrode potential of a micro-salt bridge electrode is more stable compared to a standard electrode. The implementation of this system stabilizes electrode potential and allows more precise measurements of membrane potential generated by a pH gradient. Using this system, the proton turnover rates of the mitochondrial carriers UCP1 and UCP3 are reinvestigated and compared to earlier measurements.

In electrophysiological measurements, the presence of a diffusion potential disturbs the precise measurement of the reverse potential by altering the electrode potential. Using a micro-agar salt bridge, the impact of the diffusion potential is minimized, which allows a more precise measurement of substrate turnover numbers of reconstituted recombinant membrane proteins.

組換え膜蛋白質のプロトンの回転率を分析するためのマイクロ寒天塩橋電極

To date, more than 50% of all pharmacological drugs target the transport kinetics of membrane proteins. The electrophysiological characterization of ...

A Proteoliposome-Based Efflux Assay to Determine Single-molecule Properties of Cl- Channels and Transporters

The last 15 years have been characterized by an explosion in the ability to overexpress and purify membrane proteins from prokaryotic organisms as well as from eukaryotes. This increase has been largely driven by the successful push to obtain structural information on membrane proteins. However, the ability to functionally interrogate these proteins has not advanced at the same rate and is often limited to qualitative assays of limited quantitative value, thereby limiting the mechanistic insights that they can provide. An assay to quantitatively investigate the transport activity of reconstituted Cl- channels or transporters is described. The assay is based on the measure of the efflux rate of Cl- from proteoliposomes following the addition of the K+ ionophore valinomycin to shunt the membrane potential. An ion sensitive electrode is used to follow the time-course of ion efflux from proteoliposomes reconstituted with the desired protein. The method is highly suited for mechanistic studies, as it allows for the quantitative determination of key properties of the reconstituted protein, such as its unitary transport rate, the fraction of active protein and the molecular mass of the functional unit. The assay can also be utilized to determine the effect of small molecule compounds that directly inhibit/activate the reconstituted protein, as well as to test the modulatory effects of the membrane composition or lipid-modifying reagents. Where possible, direct comparison between results obtained using this method were found to be in good agreement with those obtained using electrophysiological approaches. The technique is illustrated using CLC-ec1, a CLC-type H+/Cl- exchanger, as a model system. The efflux assay can be utilized to study any Cl- conducting channel/transporter and, with minimal changes, can be adapted to study any ion-transporting protein.

A Proteoliposome-Based Efflux Assay to Determine Single-molecule Properties of Cl- Channels and Transporters

Proteoliposomes are used to study purified channels and transporters reconstituted in a well-defined biochemical environment. An experimental procedure to measure efflux mediated by these proteins is illustrated. The steps to prepare proteoliposomes, perform the recordings, and analyze data to quantitatively determine the functional properties of the reconstituted protein are described.

プロテオリポソー​​ムベースの流出アッセイのClの単一分子特性を決定する<sup&gt; -</sup&gt;チャネルとトランスポーター

The last 15 years have been characterized by an explosion in the ability to overexpress and purify membrane proteins from prokaryotic organisms as well as ...

生物学

Measurement of Extracellular Ion Fluxes Using the Ion-selective Self-referencing Microelectrode Technique

Cells from animals, plants and single cells are enclosed by a barrier called the cell membrane that separates the cytoplasm from the outside. Cell layers such as epithelia also form a barrier that separates the inside from the outside or different compartments of multicellular organisms. A key feature of these barriers is the differential distribution of ions across cell membranes or cell layers. Two properties allow this distribution: 1) membranes and epithelia display selective permeability to specific ions; 2) ions are transported through pumps across cell membranes and cell layers. These properties play crucial roles in maintaining tissue physiology and act as signaling cues after damage, during repair, or under pathological condition. The ion-selective self-referencing microelectrode allows measurements of specific fluxes of ions such as calcium, potassium or sodium at single cell and tissue levels. The microelectrode contains an ionophore cocktail which is selectively permeable to a specific ion. The internal filling solution contains a set concentration of the ion of interest. The electric potential of the microelectrode is determined by the outside concentration of the ion. As the ion concentration varies, the potential of the microelectrode changes as a function of the log of the ion activity. When moved back and forth near a source or sink of the ion (i.e. in a concentration gradient due to ion flux) the microelectrode potential fluctuates at an amplitude proportional to the ion flux/gradient. The amplifier amplifies the microelectrode signal and the output is recorded on computer. The ion flux can then be calculated by Fick&#8217;s law of diffusion using the electrode potential fluctuation, the excursion of microelectrode, and other parameters such as the specific ion mobility. In this paper, we describe in detail the methodology to measure extracellular ion fluxes using the ion-selective self-referencing microelectrode and present some representative results.

Transporters in cell membranes allow differential segregation of ions across cell membranes or cell layers and play crucial roles during tissue physiology, repair and pathology. We describe the ion-selective self-referencing microelectrode that allows the measurement of specific ion fluxes at single cells and tissues in vivo.

イオン選択自己参照微小電極法を用いた細胞外イオンフラックスの測定

Cells from animals, plants and single cells are enclosed by a barrier called the cell membrane that separates the cytoplasm from the outside. Cell layers ...

Electrochemical Measurements of Supported Catalysts Using a Potentiostat/Galvanostat

Source: Laboratory of Dr. Yuriy Rom&#225;n &#8212; Massachusetts Institute of Technology
A potentiostat/galvanostat (often referred to as simply a potentiostat) is an instrument that measures current at an applied potential (potentiostatic operation) or measures potential at an applied current (galvanostatic operation) (Figure 1). It is the most commonly used instrument in the electrochemical characterization of anode and cathode materials for fuel cells, electrolyzers, batteries, and supercapacitors.
Conventionally, these anode and cathode materials are interfaced with a potentiostat via a three-electrode electrochemical cell. The electrode leads from the potentiostat are connected to the reference electrode, the counter electrode (often called the auxiliary electrode), and the working electrode (which contains the test material of interest). The electrochemical cell is then filled with a high ionic strength electrolyte solution, such as an acidic, alkaline, or salt solution. The media for this high ionic strength solution is typically aqueous; however, for applications necessitating higher operating cell potential windows, such as batteries and supercapacitors, non-aqueous media is often used. The cell media is degassed with an inert gas (to prevent unwanted side reactions) or with a test gas (if the test reaction involves a gas at one of the electrodes).
Alternatively, a salt bridge or membrane is employed to maintain ionic contact if the two half cells are to be measured in different electrolytes. In heterogeneous electrocatalysis, this type of &#34;two compartment&#34; cell is often used if the test molecule at the working electrode is also reactive at the counterelectrode. This happens frequently as the counterelectrode typically employed is platinum, which is a highly active catalyst for many reactions. Here, single compartment cells will be used, where all three electrodes are in the same media.
This video will explain the process of polishing a working electrode, preparing a catalyst ink, mounting the catalyst ink onto the working electrode, preparing the electrochemical cell, and then performing electrochemical measurements. The measurements that are performed include: cyclic voltammetry (CV), linear sweep voltammetry (LSV), chronopotentiometry (CP), and chronoamperometry (CA).
<img alt="Figure 1" src="/files/ftp_upload/5698/5698fig1.jpg" /> 
Figure 1. An example of a single compartment electrochemical cell. a.) Teflon cap, b.) glass cell, c.) Pt wire counter electrode, d.) working electrode, e.) Ag/AgCl reference electrode, f.) 0.5 M aqueous sulfuric acid electrolyte solution.

塩化物イオンの動的電気化学測定

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オープンパッチ・クランプのピペットで非撹拌の境界層におけるイオン濃度の測定: 流体によるイオンチャネルの制御の流れ

鉄筋コンクリート構造物から採取された検体で腐食の塩化しきい値値を決定する実験的プロトコル

薄膜白金マクロ及びマイクロ電極の電気化学粗化

偏光ゾーンの伝播を遮断するように並置されたイオン交換膜との間のイオン濃度分極のマージ

個々の電気不活性粒子の正確な電気化学的サイジング

脳組織における測定細胞外イオンの信号のための、二連と同心微小電極

組換え膜蛋白質のプロトンの回転率を分析するためのマイクロ寒天塩橋電極

プロテオリポソームベースの流出アッセイのClの単一分子特性を決定する

イオン選択自己参照微小電極法を用いた細胞外イオンフラックスの測定

ポテンショスタット/Galvanostat を使用して担持触媒の電気化学測定

塩化物イオンの動的電気化学測定

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オープン パッチ ・ クランプのピペットで非撹拌の境界層におけるイオン濃度の測定: 流体によるイオン チャネルの制御の流れ

鉄筋コンクリート構造物から採取された検体で腐食の塩化しきい値値を決定する実験的プロトコル

薄膜白金マクロ及びマイクロ電極の電気化学粗化

偏光ゾーンの伝播を遮断するように並置されたイオン交換膜との間のイオン濃度分極のマージ

個々の電気不活性粒子の正確な電気化学的サイジング

脳組織における測定細胞外イオンの信号のための、二連と同心微小電極

組換え膜蛋白質のプロトンの回転率を分析するためのマイクロ寒天塩橋電極

プロテオリポソー​​ムベースの流出アッセイのClの単一分子特性を決定する

イオン選択自己参照微小電極法を用いた細胞外イオンフラックスの測定

ポテンショスタット/Galvanostat を使用して担持触媒の電気化学測定

オープンパッチ・クランプのピペットで非撹拌の境界層におけるイオン濃度の測定: 流体によるイオンチャネルの制御の流れ

プロテオリポソームベースの流出アッセイのClの単一分子特性を決定する