Integration of microchip electrophoresis with electrochemical detection using an epoxy-based molding method to embed multiple electrode materials

This paper describes the use of epoxy-encapsulated electrodes to integrate microchip-based electrophoresis with electrochemical detection. Devices with various electrode combinations can easily be developed. This includes a palladium decoupler with a downstream working electrode material of either gold, mercury/gold, platinum, glassy carbon, or a carbon fiber bundle. Additional device components such as the platinum wires for the electrophoresis separation and the counter electrode for detection can also be integrated into the epoxy base. The effect of the decoupler configuration was studied in terms of the separation performance, detector noise, and the ability to analyze samples of a high ionic strength. The ability of both glassy carbon and carbon fiber bundle electrodes to analyze a complex mixture was demonstrated. It was also shown that a PDMS-based valving microchip can be used along with the epoxy-embedded electrodes to integrate microdialysis sampling with microchip electrophoresis and electrochemical detection, with the microdialysis tubing also being embedded in the epoxy substrate. This approach enables one to vary the detection electrode material as desired in a manner where the electrodes can be polished and modified as is done with electrochemical flow cells used in liquid chromatography.     

Carbon paste-based electrochemical detectors for microchip capillary electrophoresis/electrochemistry

The first reported use of a carbon paste electrochemical detector for microchip capillary electrophoresis (CE) is described. Poly(dimethylsiloxane) (PDMS)-based microchip CE devices were constructed by reversibly sealing a PDMS layer containing separation and injection channels to a separate PDMS layer that contained carbon paste working electrodes. End-channel amperometric detection with a single electrode was used to detect amino acids derivatized with naphthalene dicarboxaldehyde. Two electrodes were placed in series for dual electrode detection. This approach was demonstrated for the detection of copper(II) peptide complexes. A major advantage of carbon paste is that catalysts can be easily incorporated into the electrode. Carbon paste that was chemically modified with cobalt phthalocyanine was used for the detection of thiols following a CE separation. These devices illustrate the potential for an easily constructed microchip CE system with a carbon-based detector that exhibits adjustable selectivity.     

Use of micromolded carbon dual electrodes with a palladium decoupler for amperometric detection in microchip electrophoresis

The fabrication and evaluation of micromolded dual carbon ink electrodes and their integration with a fabricated palladium decoupler for use in microchip electrophoresis is described. As opposed to previous work involving carbon-based dual electrodes with microchip electrophoresis, this approach results in electrodes that are amenable to mass production in a manner where the decoupler/electrode alignment is fixed and reproducible. In this work, electrode sizes and spacings were optimized to result in dual carbon electrodes that are 1 microm in height and separated by 100 microm. Fluorescence microscopy was used to investigate leakage around the electrode/channel interface as well as to investigate what effect the dual electrodes have on band broadening phenomena. The performance of the microelectrodes was demonstrated by the separation and selective dual electrode detection of neurotransmitters in the presence of ascorbic acid. It was also found that addition of SDS to the buffer system improved both the LODs and collection efficiencies. This approach, which is the first involving carbon-based dual electrodes with an on-chip palladium decoupler, will be useful for separating and detecting neurotransmitters that are either collected by in vivo sampling or released from cells on-chip.     

Fabrication and evaluation of a carbon-based dual-electrode detector for poly(dimethylsiloxane) electrophoresis chips

The first carbon-based dual-electrode detector for microchip capillary electrophoresis (CE) is described. The poly(dimethylsiloxane) (PDMS)-based microchip CE devices were constructed by reversibly sealing a PDMS layer containing separation and injection channels to another PDMS layer containing carbon fiber working electrodes. End-channel amperometric detection was employed and the performance of the chip was evaluated using catechol. The response was found to be linear between 1 and 600 microM with an experimentally determined limit of detection (LOD) of 500 nM and a sensitivity of 30 pA/microM. Collection efficiencies for catechol ranged from 36.0 to 43.7% at field strengths of 260-615 V/cm. The selectivity that can be gained with these devices is demonstrated by the first CE-based dual-electrode detection of a Cu(II) peptide complex. These devices illustrate the potential for a rugged and easily constructed microchip CE system with an integrated carbon-based detector of similar scale.     

新型芯片电泳柱端安培检测系统的构建与评价

自行设计开发了一套便于与电泳芯片集成的一体式柱端安培检测池系统．该系统由整块透明有机玻璃精密加工而成,包括电泳芯片支架和安培检测池两部分,芯片可通过芯片插槽和不锈钢夹具固定在芯片支架上,各种检测用电极可直接通过螺母固定在安培检测池中．以100μmol／L的DA为模式分析物,分别采用直径为100、300和500μm的铂金圆盘电极与表观直径为240μm的碳纤维电极作为工作电极均在该装置上实现了良好组装和高灵敏检测．采用碳纤维工作电极对该系统的检测参数进行了优化．测试结果表明该系统在电化学清洗程序下连续六次测定100μmol／L多巴胺的峰电流相对标准偏差为3．2％,保留时间相对标准偏差为0．5％,DA的检测限为0．4μmol／L（按照S／N=3计）．该系统体积小巧,测试稳定,检测灵敏度较高,工作电极更换方便,适合作为芯片电泳柱端安培检测通用平台．     

Nanoband electrode for high-performance in-channel amperometric detection in dual-channel microchip capillary electrophoresis

A nanoband electrode detector integrated with a dual-channel polydimethylsiloxane microchip is proposed for in-channel amperometric detection in microchip capillary electrophoresis. Gold nanoband electrodes, which were fabricated on SU-8 substrates with a 100-nm-width gold layer, were introduced into the dual-channel microchip to be an electrochemical detector. Due to the nano-sized width of the detector, the noise of the amperometric detection was significantly reduced, and a high separation resolution was achieved for monitoring the analytes. The detection sensitivity of the system was improved by high signal-to-noise ratio, and a low detection limit on microchip was obtained for p-aminophenol (2.09 nM). Because of the high resolution in measuring half-peak width, the plate number that is used to evaluate the separation efficiency was 1.5-fold higher than that using 50-μm-width electrochemical detector. The effect of sample injection time and data acquisition time on separation efficiency was investigated, and an attractive separation efficiency was achieved with a plate number up to 17,500.     

Electrochemical Detection Using an Engraved Microchip – Capillary Electrophoresis Platform

The present study describes a simple strategy to integrate electrochemical detection with an assembled microchip‐capillary electrophoresis platform. The electrochemical cell was integrated with a microfluidic device consisting of five plastic squares interconnected with fused silica capillaries, forming a four‐way injection cross between the separation channel and three side‐arms (each of 15 mm in length) acting as buffer/sample reservoirs. The performance of the system was evaluated using electrodes made with either carbon ink, carbon nanotubes, or gold and under different experimental conditions of pH, capillary length, and injection time. Using this system it was possible to separate the neurotransmitters dopamine and cathecol and to quantify phenol from a real sample using a linear calibration curve with a calculated LOD of 0.7 µM. A similar concept was applied to determine glucose, by including a pre‐reactor filled with beads modified with glucose oxidase (GOx). The latter system was used to determine glucose in a commercial sample, with a recovery of 95.2 %. Overall, the presented approach represents a simple, inexpensive, and versatile approach to integrate electrochemical detection with CE separations without requiring access to microfabrication facilities.     

介电电泳芯片及其在细胞分析中的应用

简要阐述了在交流和直流电压电场中,介电电泳(DEP)芯片进行细胞分离富集的机理.按照驱动电场的差异对DEP芯片进行了分类,分析和比较了DEP芯片微电极的叉指电极、抛物线电极、堡式电极、三维电极等典型结构.特别对近年来DEP芯片在单细胞分析、细胞分离与富集以及临床细胞分析中的应用进展进行了综述,并对其应用前景和发展方向进行了展望.     

Design and development of microfabricated capillary electrophoresis devices with electrochemical detection

Our efforts have been focused on developing a self-contained and transportable microfabricated electrophoresis (CE) system with integrated electrochemical detection (ED). The current prototype includes all necessary electrodes “on-chip” and utilizes miniaturized CE and ED supporting electronics custom designed for this purpose. State-of-the-art design/modeling tools and novel microfabrication procedures were used to create recessed platinum electrodes with complex geometries and the CE/ED device from two patterned ultra-flat glass substrates. The electrodes in the bottom substrate were formed by a self-aligned etch and deposition technique followed by a photolithographic lift-off process. The microchannels (20 μm deep×65 μm wide (average)) were chemically etched into the top substrate followed by thermal bonding to complete the microchip device. CE/ED experiments were performed using 0.02 M phosphate buffer (pH 6), an analyte/buffer solution (2.2 mM dopamine, 2.3 mM catechol) and varying separation voltages (0-500 V) with a custom electronics unit interfaced to a laptop computer for data acquisition. Detection limits (S/N=3) were found to be at the micromolar level and a linear detection response was observed up to millimolar concentrations for dopamine and catechol. The microchip CE/ED system injected 50 pl volumes of sample, which corresponded to mass detection limits on the order of 200 amol. For the first time, an integrated “on-chip” multi-electrode array CE/ED device was successfully designed, fabricated and tested. The majority of the electrodes (six out of eight) in the array were capable of detecting dopamine with the amplitude of the signal (i.e., peak heights) decreasing as the electrode distance from the channel exit increased.     

Integration of a carbon microelectrode with a microfabricated palladium decoupler for use in microchip capillary electrophoresis/electrochemistry

A method to integrate a carbon microelectrode with a microfabricated palladium decoupler for use in microchip capillary electrophoresis (CE) is detailed. As opposed to previous studies with decouplers for microchip CE, the working electrode material, which is made by micromolding of a carbon ink, is different from the decoupling electrode material (palladium). The manner in which the working electrode is made does not add additional etching or lithographic steps to the fabrication of the glass electrode plate. The hybrid poly(dimethylsiloxane)/glass device was characterized with fluorescence microscopy and by monitoring the CE-based separation of dopamine. Hydrodynamic voltammograms exhibited diffusion-limited currents occurring at potentials above +1.0 V. It was also shown that the half-wave potential does not shift as the separation potential is changed, as is the case in nondecoupled systems. Gated injections of dopamine in a 25 mM boric acid buffer (pH 9.2) showed a linear response from 200 to 5 microM (r2 = 0.9992), with a sensitivity of 5.47 pA/microM and an estimated limit of detection of 2.3 microM (0.621 fmol, S/N = 3). This is the first report of coupling a carbon electrode with a decoupler in microchip CE.     

Electrochemical detection of phenolic compounds using cylindrical carbon-ink electrodes and microchip capillary electrophoresis

A simple method to fabricate cylindrical carbon electrodes for use in capillary electrophoresis (CE) microchips is described. The electrodes were fabricated using a metallic wire coated with carbon ink. Several experimental variables were studied in order to establish the best conditions to fabricate the electrode. Finally, the electrodes were integrated in a poly(dimethylsiloxane) microchip and used for the analysis of phenolic compounds. Using the optimum conditions, the analysis of a mixture of dopamine, epinephrine, catechol, and 4-aminophenol was achieved in less than 240 s, showing good linear responses (R² = 0.999) in the 0.1-190 μM range, and limits of detection (without the use of stacking or a decoupler) of 140 and 105 nM for dopamine and epinephrine, respectively.     

Simultaneous microchip enzymatic measurements of blood lactate and glucose

A miniaturized capillary electrophoretic (CE) microchip device for the simultaneous measurements of lactate and glucose is described. The new microchip bioassay protocol integrates an electrophoretic separation of lactate and glucose, post-column enzymatic reactions of these metabolites with their respective oxidase enzymes, and an amperometric (anodic) detection of enzymatically-liberated hydrogen peroxide at a gold-coated thick-film carbon detector. Factors influencing the response have been examined and optimized, and the analytical performance has been characterized. Applicability of the microchip assay to clinical samples, such as serum and blood, is demonstrated. The microchip protocol obviates cross enzymatic reactions and interferences from major oxidizable constituents common to dual glucose-lactate enzyme electrodes. Such ability to rapidly separate and quantitate lactate and glucose on a small microchip platform should find important clinical and biotechnological applications.     

Microchip capillary electrophoresis/electrochemical detection of hydrazine compounds at a cobalt phthalocyanine modified electrochemical detector

This article reports on the use of cobalt(II) phthalocyanine (CoPc)-modified carbon paste amperometric detector for monitoring hydrazine compounds following their microchip separation. The marked catalytic electrochemical properties of CoPc-modified electrode display enhanced sensitivity compared with unmodified carbon pastes at a relatively low detection potential (+0.5 V versus Ag/AgCl). Factors influencing the on-chip separation and detection processes have been optimized. Three hydrazines (hydrazine, 1,1 dimethylhydrazine, and phenylhydrazine) have been separated within 130 s at a separation voltage of 1 kV using a 10 mM phosphate run buffer (pH 6.5). The detection limits obtained from using the CoPc-modified carbon paste electrodes for hydrazine and phenylhydrazine are 0.5 and 0.7 μM, respectively, with linearity over the 20–200 μM range examined. Such miniaturization and speed advantages of microchip CE are coupled to the highly sensitivity and convenient preparation of CoPc-modified carbon paste electrode. The resulting microsystem should be attractive for field monitoring of toxic hydrazine compounds in environmental applications.     

Screen‐Printed Contactless Conductivity Detector for Microchip Capillary Electrophoresis

A new method for mass fabrication of silver ink conductivity detector electrodes for poly(methylmethacrylate) (PMMA) microchip electrophoretic systems has been developed based on screen‐printing technology. Printing of silver conductivity electrodes was performed through a patterned stencil on thin PMMA sheets. Following the electrode fabrication, the PMMA sheets are cut into cover sheets, and are aligned and sealed to the channel plate thus establishing a complete microchip separation device. The effects of the electrode width and spacing on the response and resolution have been investigated and the optimized electrode performance was compared to commonly used aluminum electrodes in the determination of ammonium, methyl ammonium, and sodium. The utility of the screen‐printed contactless conductivity detector (SPCCD) electrodes is further demonstrated for the separation and detection of organic acids with excellent reproducibility (RSD values of 3.7% and 4.1% for oxalate and tartrate, respectively). The thick‐film fabrication of the electrode material demonstrates the ability to mass‐fabricate detection devices with total process of device fabrication requiring less than 4 h (including the fabrication of channel plate, cover sheet with the electrodes, and subsequent bonding). The fabrication method described here is convenient and does not compromise the detector performance, hence offers great promise for producing single use field deployable analytical microsystems.     

Performance evaluation of a capillary electrophoresis electrochemical chip integrated with gold nanoelectrode ensemble working and decoupler electrodes

This paper presents a capillary electrophoresis poly(methyl methacrylate) (PMMA) based microchip for electrochemical detection applications featuring embedded gold nanoelectrode ensemble (GNEE) working and decoupler electrodes. In fabricating the microchip, the GNEE films are pressed directly onto the metallic electrode structures using a hot embossing technique, and the microfluidic channels are then sealed using a low-temperature azeotropic solvent bonding method. The detection performance of the microchip is evaluated using dopamine and catechol analytes for illustration purposes. The experimental results show that the GNEE working electrode provides a significantly higher signal response than that obtained from a bulk gold electrode when applied to the detection of dopamine analyte. Compared to a conventional bulk palladium decoupler electrode, the GNEE decoupler electrode reduces both the amplitude of the charge current (3.5 nA vs. 18.7 nA) and the baseline drift at higher separation voltages. The measured baseline current drift for the microchip equipped the proposed GNEE decoupler electrode is around three times smaller than the microchip with the palladium decoupler electrode under the applied separation electric field from 40 V/cm to 240 V/cm. Finally, when detecting a mixture of 1mM dopamine and 1mM catechol, the calculated signal response of the microchip with a GNEE decoupler electrode is approximately five times higher than that obtained from a microchip with a bulk Pd decoupler electrode, resulting in the detection limit of 1 microM for the proposed GNEE-based microchip device. Overall, the results indicate that the proposed capillary electrophoresis-electrochemical detection (CE-ED) microchip with embedded GNEE working and decoupler electrodes provides an ideal solution for sample detection in lab-on-a-chip and micro total analysis applications.     

Fabrication of SU-8 based microchip electrophoresis with integrated electrochemical detection for neurotransmitters

A new SU-8 based microchip capillary electrophoresis (MCE) device has been developed for the first time with integrated electrochemical detection. Embedded electrophoretic microchannels have been fabricated with a multilayer technology based on bonding and releasing steps of stacked SU-8 films. This technology has allowed the monolithic integration in the device of the electrochemical detection system based on platinum electrodes. The fabrication of the chips presented in this work is totally compatible with reel-to-reel techniques, which guarantee a low cost and high reliability production. The influence of relevant experimental variables, such as the separation voltage and detection potential, has been studied on the SU-8 microchip with an attractive analytical performance. Thus, the effective electrical isolation of the end-channel amperometric detector has been also demonstrated. The good performance of the SU-8 device has been proven for separation and detection of the neurotransmitters, dopamine (DA) and epinephrine (EP). High efficiency (30,000-80,000 N/m), excellent precision, good detection limit (450 nM) and resolution (0.90-1.30) has been achieved on the SU-8 microchip. These SU-8 devices have shown a better performance than commercial Topas (thermoplastic olefin polymer of amorphous structure) microchips. The low cost and versatile SU-8 microchip with integrated platinum film electrochemical detector holds great promise for high-volume production of disposable microfluidic analytical devices.     

芯片毛细管电泳-安培检测系统

由于安培检测具有的高灵敏度、低成本、低能耗、易集成化便携化、与微加工技术匹配等特点,芯片毛细管电泳-安培检测系统（μCE-AD）的研究近年来得到人们广泛的关注。本文结合本课题组的研究工作,对近年来μCE-AD的研究进展进行评述;重点讨论了近年来在芯片的设计、集成化电极的制备、消除分离电压的干扰等方面的进展;同时介绍了利用分离电场拓展检测范围、阵列电极和阵列通道、化学修饰电极的应用、新型进样技术和试样预处理等方面的新成就;最后展望了未来μCE-AD的发展趋势。     

Detecting thiols in a microchip device using micromolded carbon ink electrodes modified with cobalt phthalocyanine

This paper describes the fabrication and evaluation of a chemically modified carbon ink microelectrode to detect thiols of biological interest. The detection of thiols, such as homocysteine and cysteine, is necessary to monitor various disease states. The biological implications of these thiols generate the need for miniaturized detection systems that enable portable monitoring as well as quantitative results. In this work, we utilize a microchip device that incorporates a micromolded carbon ink electrode modified with cobalt phthalocyanine to detect thiols. Cobalt phthalocyanine (CoPC) is an electrocatalyst that lowers the potential needed for the oxidation of thiols. The CoPC/carbon ink composition was optimized for the micromolding method and the resulting microelectrode was characterized with microchip-based flow injection analysis. It was found that CoPC lowers the overpotential for thiols but, as compared to direct amperometric detection, a pulsed detection scheme was needed to constantly regenerate the electrocatalyst surface, leading to improved peak reproducibility and limits of detection. Using the pulsed method, cysteine exhibited a linear response between 10-250 microM (r(2) = 0.9991) with a limit of detection (S/N = 3) of 7.5 microM, while homocysteine exhibited a linear response between 10-500 microM (r(2) = 0.9967) with a limit of detection of 6.9 microM. Finally, to demonstrate the ability to measure thiols in a biological sample using a microchip device, the CoPC-modified microelectrode was utilized for the detection of cysteine in the presence of rabbit erythrocytes.     

A simple PDMS-based electro-fluidic interface for microchip electrophoretic separations

High voltage electrodes for electrophoresis have been integrated into a polymer layer that can be reversibly bound to glass microchips for electrophoretic separations. By using the liquid precursor to the polymer polydimethylsiloxane (PDMS), platinum electrodes and reservoirs can be positioned prior to solidification, providing a simple and flexible method for electrode interface construction. Field strengths up to 875 V cm(-1) over an 8 cm separation channel can be applied to the system without any loss in performance of the interface. The interface can function as an electro-fluidic interface between the high voltage power supply and the separation channel and, when reversibly sealed to an etched glass plate, functions as a cover plate establishing a hybrid PDMS-glass microchip in which the electrodes are directly integrated onto the device. The versatility of this approach is not only demonstrated by separating DNA fragments in a novel buffer sieving matrix, but also with the molecular diagnostic analysis of a variety of DNA samples for Duschenne Muscular Dystrophy and cytomegalovirus (CMV) infection, using both microchip interface configurations.     

Convenient enantioseparation by monolithic imprinted capillary clamped in a chip with electrochemical detection

A microchip integrated with a monolithic imprinted capillary has been manufactured for performing the chip-based capillary electrochromatographic enantioseparation. The microporous monolith anchored on the inner wall of the microchannel was prepared by in situ chemical copolymerization, and characterized with scanning electron microscopy, IR spectroscopy, and solid-state UV-vis spectroscopy. The monolithic network with high porosity gave a large surface area, good permeability, low mass-transfer resistance, and thus high separation efficiency. A portable microchip was conveniently constructed by integrating an imprinted capillary with 5-cm length as the separation channel and a carbon fiber microdisk working electrode for amperometric detection. Using L-tyrosine (L-Tyr) as the template molecule, Tyr enantiomers could be baseline separated within 55 s under the optimized preparation and separation conditions. The linear ranges for online amperometric detection of both Tyr enantiomers were from 20 to 2400 μM. The microporous monolithic chip strategy exhibited excellent separation efficiency and promising analytical application in enantioseparation. It opens an avenue for high-throughput screening of chiral compounds.