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.     

Fabrication and evaluation of single- and dual-channel (Pi-design) microchip electrophoresis with electrochemical detection

A new dual-channel microchip capillary electrophoresis (MCE) has been developed on glass substrates for the first time with electrochemical detection. Dual-channel (called Pi-design) as well as single-channel microchips have been fabricated on soda-lime glass using photolithography, wet etching and thermal bonding. Moreover, a laser writing system has been applied for the fabrication of photomasks with the different microchip designs (single- and dual-channel configurations). The microfabricated channels have been characterized by optical, confocal and scanning electron microscopy. The resulting single- and dual-channel microchips have been evaluated using an end-channel amperometric detector based on one (single-channel) or two (dual-channel) 100-mum gold wires aligned at the outlet of the separation channel. Parameters affecting the separation of several phenolic compounds (dopamine, p-aminophenol and hydroquinone) have been studied in the glass microchips. Thus, the influence of separation voltage, detection potential and background electrolyte has been examined in the single-channel microchip. Different total length microchannel has been compared. Furthermore, the possibility of carrying out two simultaneous measurements has been demonstrated in the new dual-channel microchip electrophoresis. The injection format has been checked and resulted to be critical, in such a way that a special and new form is employed for obtaining simultaneous signals at both channels. Analytical characteristics, such as sensitivity and reproducibility have been evaluated and resulted very adequate.     

Microchip reversed-phase liquid chromatography with packed column and electrochemical flow cell using polystyrene/poly(dimethylsiloxane)

A microchip pressure-driven liquid chromatography (LC) with a packed column and an electrochemical flow cell has been developed by using polystyrene (PS) and poly(dimethylsiloxane) (PDMS). The cylindrical separation column with packed octadecyl silica particles was fabricated in the PS substrate. The three electrode system (working, reference, and counter electrode) for amperometric detection was fabricated onto the PS substrate, using the Au deposition, photolithography, and chemical etching. The detector flow cell was formed by sealing the electrode system with a PDMS chip containing a channel. In this flow cell, the effect of working electrode width (in the direction of flow) on chromatographic parameters, such as peak width and peak resolution were studied in electrode width ranging 50-5,000 microm. The effect of electrode width on sensitivity (current intensity, current density, and S/N ratio) was also examined. The sensitivity was discussed by simulating the concentration profile generated around the working electrode. The effects of the column packing size and the column size on the separation efficiency were examined. In this study, a good separation of three catechins was successfully achieved and the detection limits for (+)-catechin, epicatechin, and epigallocatechin gallate were 350, 450, and 160 nM, respectively.     

Preparation of metal nanoband microelectrode on poly(dimethylsiloxane) for chip-based amperometric detection

We proposed herein a novel approach for fabricating nanoband microelectrodes for electrochemical detection on an electrophoresis microchip. The metal films were first obtained via region-selective electroless deposition of gold or copper films on PDMS substrates by selective region plasma oxidation through shadow masking. Both metal films show uniform surfaces with the thickness at the level of 100 nm. By casting another PDMS layer on the metal films, the cross section of the sandwich structures can be used as nanoband microelectrodes, which can be renewed just by cutting. These nanoband microelectrodes are successfully used as electrochemical detectors in microchip electrophoresis for the detection of amino acids, proteins and neurotransmitter molecules. Moreover, integrating an Au-Cu double-metal detector with a double-channel electrophoresis system, we can easily distinguish electroactive amino acids from that of non-electroactive amino acids.     

Poly(methylmethacrylate) and Topas capillary electrophoresis microchip performance with electrochemical detection

A capillary electrophoresis (CE) microchip made of a new and promising polymeric material: Topas (thermoplastic olefin polymer of amorphous structure), a cyclic olefin copolymer with high chemical resistance, has been tested for the first time with analytical purposes, employing an electrochemical detection. A simple end-channel platinum amperometric detector has been designed, checked, and optimized in a poly-(methylmethacrylate) (PMMA) CE microchip. The end-channel design is based on a platinum wire manually aligned at the exit of the separation channel. This is a simple and durable detection in which the working electrode is not pretreated. H(2)O(2) was employed as model analyte to study the performance of the PMMA microchip and the detector. Factors influencing migration and detection processes were examined and optimized. Separation of H(2)O(2) and L-ascorbic acid (AsA) was developed in order to evaluate the efficiency of microchips using different buffer systems. This detection has been checked for the first time with a microchip made of Topas, obtaining a good linear relationship for mixtures of H(2)O(2) and AsA in different buffers.     

Amperometric detector designs for capillary electrophoresis microchips

Electrochemical (EC) detection is a sensitive and miniaturisable detection mode for capillary electrophoresis (CE) microchips. Detection cell design is very important in order to ensure electrical isolation from the high separation voltage. Amperometric detectors with different designs have been developed for coupling EC detection to CE-microchips. Different working electrode alignment: in-channel or end-channel has been tested in conjunction with several materials: gold, platinum or carbon. The end-channel detector was based on a platinum or gold wire manually aligned at the exit of the separation channel. Thick- (screen-printed carbon electrode) and thin-film (sputtered gold film) electrodes have also been employed with this configuration, but with a different design that allowed the rapid replacement of the electrode. The in-channel detector was based on a gold film within the separation channel. A gold-based dual electrode detector, which combined for the first time in- and end-channel detection, has been also tested. These amperometric detectors have been evaluated in combination to poly(methylmethacrylate) (PMMA) and Topas (thermoplastic olefin polymer of amorphous structure) CE-microchips. Topas is a new and promising cyclic olefin copolymer with high chemical resistance. Relevant parameters of the polymer microchip separation such as precision, efficiency or resolution and amperometric detection were studied with the different detector designs using p-aminophenol and L-ascorbic acid as model analytes in Tris-based buffer pH 9.0.     

Microchip electrophoresis with wall-jet electrochemical detector: influence of detection potential upon resolution of solutes

This report studies the electrochemical response of wall-jet detector for microchip electrophoresis (microCE). It shows that in wall-jet configuration, the electrochemical detector operates in coulometric mode and that there is an influence of detection potential upon peak width and therefore upon the resolution of solutes. Upon raising the detection potential from +0.3 to +0.9 V, the resolution between model analytes, dopamine and catechol, increases from 0.63 to 2.90. The reasons for this behavior originate in wall-jet detector design and in its typically significant higher detector volume than the volume of injected sample. The conversion efficiency of the wall-jet electrochemical detection cell was found to be 97.4% for dopamine and 98.0% for catechol. The paper brings deeper understanding of operations of wall-jet electrochemical detectors for microchip devices, and it explains previously reported significantly sharper peaks when electrocatalytic electrodes (i.e., palladium and carbon nanotube) were used in microCE-electrochemistry wall-jet detector.     

Nickel Amperometric Detector Prepared by Electroless Deposition for Microchip Electrophoretic Measurement of Alcohols and Sugars

Nickel was deposited directly at the end of an electrophoretic glass microchip using an electroless deposition procedure leading to an attractive amperometric detector for sugars and alcohols. Such direct electroless deposition of nickel at the end of the separation chip greatly simplifies the preparation of on‐chip electrochemical detectors. Variables affecting the preparation and operation of the new detector are characterized and optimized. The integrated capillary electrophoresis‐electrochemical detection microsystem was evaluated for the separation of alcohols and sugars in connection to assays of different beer and wine samples. Linear calibration plots and favorable detection limits are found that meet the needs of real sample (wine and beer) analyses. This represents the first example of using nickel electrode and detecting alcohols on microchip platforms.     

Recent developments in amperometric detection for microchip capillary electrophoresis

The interest in microfluidic devices has increased considerably over the past decade due to the numerous advantages of working within a miniature, microfabricated format. This review focuses on recent advances in coupling amperometric detection with microchip capillary electrophoresis (CE). Advances in electrochemical cell design, isolation of the detector from the separation field, and integration of both pre- and postseparation reaction chambers are discussed. The use of microchip CE with amperometric detection for enzyme/immunoassays, clinical and environmental assays, and the determination of neurotransmitters is described.     

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.     

In-channel modification of electrochemical detector for the detection of bio-targets on microchip

The coulometric efficiency (C_eff) of an amperometric detector integrated on PDMS/glass capillary electrophoresis microfluidic device (microchip) has been enhanced by in-channel electrochemical modification. In-channel electrochemical deposition of gold particles was performed in order to vertically increase the surface area of the Au sensing microelectrode. The roughness of the electrodes was characterized using scanning electron microscopy and profilometric analysis. The degree of electrode modification was also characterized by roughness factor determination. Separation processes including detection potential was optimized and the analytical performance of the microchip was tested using a mixture of dopamine (DA) and catechol (CA). The modified electrochemical detector provided well-resolved separation of DA and CA in less than 60 s with enhanced sensitivity; no peak broadening was observed. The limit of detection using in-channel modification of working electrode for DA and CA are 60 and 110 nM, respectively. Thus, in-channel electrochemical deposition of metallic particles should be used to enhance the C_eff of integrated amperometric detection of analytes with good redox properties in order to obtain lower LODs.     

Modified Hadamard transform microchip electrophoresis

Sensitivity is a crucial point in the development applications for medicine or environmental samples in which the analytes are present in the nanomolar range. Besides further technical development of detection systems, the multiplex sample injection technique can be applied for enhancing the signal-to-noise ratio. Hadamard transform is easily applied to microchip electrophoresis due to the fact that sample injection is generally achieved through cross, double-tee, or tee injector structures. This paper reports the first demonstration of a modified Hadamard transform electrophoresis on a microchip by using an amperometric detector. Contrary to the previous Hadamard applications, the resolution (number of points per unit of time) of electropherograms obtained is independent of the number of injections.     

Carbon fiber nanoelectrodes applied to microchip electrophoresis amperometric detection of neurotransmitter dopamine in rat pheochromocytoma (PC12) cells

Microelectrodes have been adopted in electrochemical detection for CE or microchip CE in recent years. In this paper, the use of nanoelectrodes (with tip diameter of 100-300 nm) as the electrochemical detector in microchip CE is firstly reported. The experimental results indicated that both the sensitivity and resolution of microchip CE with the carbon fiber nanoelectrode (CFNE) amperometric detection have been improved markedly comparing with the traditional microelectrodes. The detection limit of dopamine (S/N = 3) is 5.9x10(-8) M, which is one or two orders of magnitude lower than that reported so far, and the resolution of dopamine (DA) and isoprenaline (IP) has also improved from 0.6 (using 7 mum carbon fiber microelectrodes, CFME) to 1.0. We assembled a novel and easily operated microchip CE system with end-column amperometric detection, which allows the convenient and fast replacement of the passivated electrodes. Under the optimized condition, the RSDs of peak height and migration time are 1.47 and 0.31%, respectively (n = 40), indicating that the system displays excellent reproducibility. The nanoelectrode-based microchip CE system has been successfully applied to the determination of DA in cultured rat pheochromocytoma (PC12) cells, and the average content of DA in an individual PC12 cell is 0.54 +/- 0.07 fmol, which is in good agreement with that reported in the literature.     

In-channel amperometric detection for microchip electrophoresis using a wireless isolated potentiostat

The combination of microchip electrophoresis with amperometric detection leads to a number of analytical challenges that are associated with isolating the detector from the high voltages used for the separation. While methods such as end-channel alignment and the use of decouplers have been employed, they have limitations. A less common method has been to utilize an electrically isolated potentiostat. This approach allows placement of the working electrode directly in the separation channel without using a decoupler. This paper explores the use of microchip electrophoresis and electrochemical detection with an electrically isolated potentiostat for the separation and in-channel detection of several biologically important anions. The separation employed negative polarity voltages and tetradecyltrimethylammonium bromide (as a buffer modifier) for the separation of nitrite (NO??), glutathione, ascorbic acid, and tyrosine. A half-wave potential shift of approximately negative 500 mV was observed for NO?? and H?O? standards in the in-channel configuration compared to end-channel. Higher separation efficiencies were observed for both NO?? and H?O? with the in-channel detection configuration. The limits of detection were approximately two-fold lower and the sensitivity was approximately two-fold higher for in-channel detection of nitrite when compared to end-channel. The application of this microfluidic device for the separation and detection of biomarkers related to oxidative stress is described.     

Microchannel-electrode alignment and separation parameters comparison in microchip capillary electrophoresis by scanning electrochemical microscopy

The end of separation channel in a microchip was electrochemically mapped using the feedback imaging mode of scanning electrochemical microscopy (SECM). This method provides a convenient way for microchannel-electrode alignment in microchip capillary electrophoresis. Influence of electrode-to-channel positions on separation parameters in this capillary electrophoresis-electrochemical detection (CE-ED) was then investigated. For the trapezoid shaped microchannel, detection in the central area resulted in the best apparent separation efficiency and peak shape. In the electrode-to-channel distance ranging from 65 to 15mum, the limiting peak currents of dopamine increased with the decrease of the detection distance due to the limited diffusion and convection of the sample band. Results showed that radial position and axial distance of the detection electrode to microchannel was important for the improvement of separation parameters in CE amperometric detection.     

A high-performance polycarbonate electrophoresis microchip with integrated three-electrode system for end-channel amperometric detection

A fully integrated polycarbonate (PC) microchip for CE with end-channel electrochemical detection operated in an amperometric mode (CE-ED) has been developed. The on-chip integrated three-electrode system consisted of a gold working electrode, an Ag/AgCl reference electrode and a platinum counter electrode, which was fabricated by photo-directed electroless plating combined with electroplating. The working electrode was positioned against the separation channel exit to reduce post-channel band broadening. The electrophoresis high-voltage (HV) interference with the amperometric detection was assessed with respect to detection noise and potential shifts at various working-to-reference electrode spacing. It was observed that the electrophoresis HV interference caused by positioning the working electrode against the channel exit could be diminished by using an on-chip integrated reference electrode that was positioned in close proximity (100 microm) to the working electrode. The CE-ED microchip was demonstrated for the separation of model analytes, including dopamine (DA) and catechol (CA). Detection limits of 132 and 164 nM were achieved for DA and CA, respectively, and a theoretical plate number of 2.5x10(4)/m was obtained for DA. Relative standard deviations in peak heights observed for five runs of a standard solution containing the two analytes (0.1 mM for each) were 1.2 and 3.1% for DA and CA, respectively. The chip could be continuously used for more than 8 h without significant deterioration in analytical performance.     

Miniaturized thin-layer radial flow cell with interdigitated ring-shaped microarray electrode used as amperometric detector for capillary electrophoresis

A chip-type thin-layer radial flow cell was developed as an amperometric detector for capillary electrophoresis. We fabricated a carbon film-based interdigitated ring-shaped array (IDRA) microelectrode with a 2 microm bandwidth and an almost 1 microm gap on a glass plate and used it as a working electrode. A fused-silica capillary was arranged above the IDRA electrode using a guide hole drilled through the acryl plate that formed the flow cell lid. A flow channel for use in connecting the outlet capillary was also fabricated in the acryl plate. We characterized the analytical performance of the IDRA electrode in the microchip flow cell in terms of linear concentration range, sensitivity and concentration detection limit. We achieved a collection efficiency and catechol redox cycle at the IDRA microelectrode of 65% and 1.71, respectively, and thus a high sensitivity and low detection limit of 392.9 pA/microM and 15 nM for dopamine hydrochloride. We examined the reproducibility of the detector and found that the run-to-run and detector-to-detector relative standard deviations were both less than 10%.     

Use of a Carbon-ink Microelectrode Array for Signal Enhancement in Microchip Electrophoresis with Electrochemical Detection

In this communication, we demonstrate that a carbon ink microelectrode array, where the electrodes are held at the same potential, affords significant signal enhancement in microchip electrophoresis with amperometric detection. The ability to fabricate an array of carbon ink microelectrodes with a palladium decoupler was demonstrated and the resulting electrodes were integrated with a valving microchip design. The use of an 8 electrode array led to a significant improvement in the limits of detection at the expense of separation resolution due to the increased detection zone size. It is also shown that microdialysis sampling can be integrated with the microchip device and a multi-analyte separation achieved.     

芯片毛细管电泳电化学检测

评述了芯片毛细管电泳各种电化学检测尤其是安培检测中工作模式、工作电极、分离电流的消除、应用等方面的进展,并进行了展望。     

An end-channel amperometric detector for microchip capillary electrophoresis

An end-channel amperometric detector with a guide tube for working electrode was designed and integrated on a home-made glass microchip. The guide tube was directly patterned and fabricated at the end of the detection reservoir, which made the fixation and alignment of working electrode relatively easy. The fabrication was carried out in a two-step etching process. A 30 μm carbon fiber microdisk electrode and Pt cathode were also integrated onto the amperometric detector. The characteristics and primary performance of the home-made microchip capillary electrophoresis (MCCE) were investigated with neurotransmitters. The baseline separation of dopamine (DA), catechol (CA) and epinephrine (EP) was achieved within 80 s. Separation parameters such as injection time, buffer components, pH of the buffer were studied. Relative standard deviations of not more than 6.0% were obtained for both peak currents and migration times. Under the selected separation conditions, the response for DA was linear from 5 to 200 μM and from 20 to 800 μM for CA. The limits of detection of DA and CA were 0.51 and 2.9 μM, respectively (S/N=3).