A microchip electrophoresis system with integrated in-plane electrodes for contactless conductivity detection

We present a new approach for contactless conductivity detection for microchip-based capillary electrophoresis (CE). The detector integrates easily with well-known microfabrication techniques for glass-based microfluidic devices. Platinum electrodes are structured in recesses in-plane with the microchannel network after glass etching, which allows precise positioning and batch fabrication of the electrodes. A thin glass wall of 10-15 microm separates the electrodes and the buffer electrolyte in the separation channel to achieve the electrical insulation necessary for contactless operation. The effective separation length is 34 mm, with a channel width of 50 microm and depth of 12 microm. Microchip CE devices with conductivity detection were characterized in terms of sensitivity and linearity of response, and were tested using samples containing up to three small cations. The limit of detection for K+ (18 microM) is good, though an order of magnitude higher than for comparable capillary-based systems and one recently reported example of contactless conductivity on chip. However, an integrated field-amplified stacking step could be employed prior to CE to preconcentrate the sample ions by a factor of four.     

Fabrication and characterization of semicircular detection electrodes for contactless conductivity detector - CE microchips

This study uses simple and reliable microfabrication techniques to fabricate CE biochips, integrating a novel contactless conductivity detector in a miniaturized detection system in a microfluidic biochip. The off-channel electrodes are deposited around side channels by Au sputtering and patterned using a standard "lift-off" process. A vacuum fusion bonding process is employed to seal the lower substrate containing the microchannels and the electrodes to an upper glass cover plate. The variations in the capacitance between the semicircular detection electrodes in the side channels are measured as different samples and ions pass through the detection region of the CE separation channel. Samples of Rhodamine B, commercial sports drinks, mineral waters, and a red wine, respectively, are mixed in different buffer solutions, separated, and successfully detected using the developed device. The semicircular detection electrodes for the contactless conductivity detector have microscale dimensions and provide a valuable contribution to the realization of the lab-on-a-chip concept.     

Conductivity detection for conventional and miniaturised capillary electrophoresis systems

Since the introduction of capillary electrophoresis (CE), conductivity detection has been an attractive means of detection. No additional chemical properties are required for detection, and no loss in sensitivity is expected when miniaturising the detector to scale with narrow-bore capillaries or even to the microchip format. Integration of conductivity and CE, however, involves a challenging combination of engineering issues. In conductivity detection the resistance of the solution is most frequently measured in an alternating current (AC) circuit. The influence of capacitors both in series and in parallel with the solution resistance should be minimised during conductivity measurements. For contact conductivity measurements, the positioning and alignment of the detection electrodes is crucial. A contact conductivity detector for CE has been commercially available, but was withdrawn from the market. Microfabrication technology enables integration and precise alignment of electrodes, resulting in the popularity of conductivity detection in microfluidic devices. In contactless conductivity detection, the alignment of the electrodes with respect to the capillary is less crucial. Contactless conductivity detection (CCD) was introduced in capillary CE, and similar electronics have been applied for CCD using planar electrodes in microfluidic devices. A contactless conductivity detector for capillaries has been commercialised recently. In this review, different approaches towards conductivity detection in capillaries and chip-based CE are discussed. In contrast to previous reviews, the focus of the present review is on the technological developments and challenges in conductivity detection in CE.     

A chip-based capillary electrophoresis-contactless conductivity microsystem for fast measurements of low-explosive ionic components

A miniaturized analytical system for separating and detecting inorganic explosive residues, based on the coupling of a micromachined capillary electrophoresis (CE) chip with a contactless conductivity detector is described. The low electroosmotic flow (EOF) of the poly(methylmethacrylate) (PMMA) chip material facilitates the rapid switching between analyses of cations and anions using the same microchannel and run buffer (and without an EOF modifier), and hence offers rapid (< 1 min) measurement of seven explosive-related cations and anions. Experimental parameters relevant to the separation and detection processes have been optimized. Addition of a 18-crown-6 ether modifier has been used for separating the peaks of co-migrating potassium and ammonium ions. The ionic-explosive microchip system combines the distinct advantages of contactless conductivity detection with the attractive features of plastic CE microchips. The new microsystem offers great promise for monitoring explosive-related ions at the sample source, with significant advantages of speed/warning, efficiency, cost, or sample size.     

A Glass/PDMS Hybrid Microfluidic Chip Embedded with Integrated Electrodes for Contactless Conductivity Detection

A new glass/PDMS hybrid chip for contactless conductivity detection is reported. This chip consists of a glass substrate with microchannels and a PDMS cover sheet embedded with a small integrated electrode plate. In the region of detection, electrodes are insulated from the microchannel by a formed PDMS membrane about 100 μm in thickness. Without any modification, this glass/PDMS chip is suitable for contactless conductivity detection with good properties, such as excellent heat-dissipation, stable electroosmotic flow, high separation efficiency, satisfactory sensitivity, simple construction and high degree of integration. Its feasibility and performance had been demonstrated by analyzing inorganic ions and amino acids in mixtures, and alkaloids in traditional Chinese medicine. The limits of detection reached micromole per liter (μmol L^?1) levels. This microchip could be promising for mass production due to its stability, reproducibility, ease of fabrication and low cost.     

Separation of non-UV-absorbing synthetic polyelectrolytes by CE with contactless conductivity detection

A specific method for the separation and detection of non-UV-absorbing polyelectrolytes has been developed. The analysis of such polyelectrolytes by liquid chromatography is nearly impossible due to strong ionic interactions and charge density effects. CE makes use of these charge density effects and thus enables for proper separation. A capacitively coupled contactless conductivity detector has been applied for the detection in CE. A low molar mass poly(acrylic acid) sodium salt standard (PAA1.3k) was separated in free solution CE and detected with the contactless conductivity detector. Different amphoteric electrolytes have been tested for their applicability as BGE for the separation of polyelectrolytes with conductivity detection. It has been shown that the best detection results are obtained with an arginine-sorbate buffer.     

A three-layer PMMA electrophoresis microchip with Pt microelectrodes insulated by a thin film for contactless conductivity detection

A three-layer poly (methyl methacrylate) (PMMA) electrophoresis microchip integrated with Pt microelectrodes for contactless conductivity detection is presented. A 50 μm-thick PMMA film is used as the insulating layer and placed between the channel plate (containing the microchannel) and the electrode plate (containing the microelectrode). The three-layer structure facilitates the achievement of a thin insulating layer, obviates the difficulty of integrating microelectrodes on a thin film, and does not compromise the integration of microchips. To overcome the thermal and chemical incompatibilities of polymers and photolithographic techniques, a modified lift-off process was developed to integrate Pt microelectrodes onto the PMMA substrate. A novel two-step bonding method was created to assemble the complete PMMA microchip. A low limit of detection of 1.25 μg ml(-1) for Na(+) and high separation efficiency of 77,000 and 48,000 plates/m for Na(+) and K(+) were obtained when operating the detector at a low excitation frequency of 60 kHz.     

A Glass/PDMS Hybrid Microfluidic Chip Embedded with Integrated Electrodes for Contactless Conductivity Detection

A new glass/PDMS hybrid chip for contactless conductivity detection is reported. This chip consists of a glass substrate with microchannels and a PDMS cover sheet embedded with a small integrated electrode plate. In the region of detection, electrodes are insulated from the microchannel by a formed PDMS membrane about 100 μm in thickness. Without any modification, this glass/PDMS chip is suitable for contactless conductivity detection with good properties, such as excellent heat-dissipation, stable electroosmotic flow, high separation efficiency, satisfactory sensitivity, simple construction and high degree of integration. Its feasibility and performance had been demonstrated by analyzing inorganic ions and amino acids in mixtures, and alkaloids in traditional Chinese medicine. The limits of detection reached micromole per liter (μmol L⁻¹) levels. This microchip could be promising for mass production due to its stability, reproducibility, ease of fabrication and low cost.

     

Integration of electrochemistry in micro-total analysis systems for biochemical assays: Recent developments

Micro-total analysis systems (μTAS) integrate different analytical operations like sample preparation, separation and detection into a single microfabricated device. With the outstanding advantages of low cost, satisfactory analytical efficiency and flexibility in design, highly integrated and miniaturized devices from the concept of μTAS have gained widespread applications, especially in biochemical assays. Electrochemistry is shown to be quite compatible with microanalytical systems for biochemical assays, because of its attractive merits such as simplicity, rapidity, high sensitivity, reduced power consumption, and sample/reagent economy. This review presents recent developments in the integration of electrochemistry in microdevices for biochemical assays. Ingenious microelectrode design and fabrication methods, and versatility of electrochemical techniques are involved. Practical applications of such integrated microsystem in biochemical assays are focused on in situ analysis, point-of-care testing and portable devices. Electrochemical techniques are apparently suited to microsystems, since easy microfabrication of electrochemical elements and a high degree of integration with multi-analytical functions can be achieved at low cost. Such integrated microsystems will play an increasingly important role for analysis of small volume biochemical samples. Work is in progress toward new microdevice design and applications.     

Hot embossing of electrophoresis microchannels in PMMA substrates using electric heating wires

A simple method based on electric heating wires has been developed for the rapid fabrication of poly(methyl methacrylate) (PMMA) electrophoresis microchips in ordinary laboratories without the need for microfabrication facilities. A piece of stretched electric heating wire placed across the length of a PMMA plate along its midline was sandwiched between two microscope slides under pressure. Subsequently, alternating current was allowed to pass through the wire to generate heat to emboss a separation microchannel on the PMMA separation channel plate at room temperature. The injection channel was fabricated using the same procedure on a PMMA sheet that was perpendicular to the separation channel. The complete microchip was obtained by bonding the separation channel plate to the injection channel sheet, sealing the channels inside. The electric heating wires used in this work not only generated heat; they also served as templates for embossing the microchannels. The prepared microfluidic microchips have been successfully employed in the electrophoresis separation and detection of ions in connection with contactless conductivity detection.     

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.     

Novel microfluidic device for the continuous separation of cancer cells using dielectrophoresis

We describe the design, microfabrication, and testing of a microfluidic device for the separation of cancer cells based on dielectrophoresis. Cancer cells, specifically green fluorescent protein‐labeled MDA‐MB‐231, are successfully separated from a heterogeneous mixture of the same and normal blood cells. MDA‐MB‐231 cancer cells are separated with an accuracy that enables precise detection and counting of circulating tumor cells present among normal blood cells. The separation is performed using a set of planar interdigitated transducer electrodes that are deposited on the surface of a glass wafer and slightly protrude into the separation microchannel at one side. The device includes two parts, namely, a glass wafer and polydimethylsiloxane element. The device is fabricated using standard microfabrication techniques. All experiments are conducted with low conductivity sucrose‐dextrose isotonic medium. The variation in response between MDA‐MB‐231 cancer cells and normal cells to a certain band of alternating‐current frequencies is used for continuous separation of cells. The fabrication of the microfluidic device, preparation of cells and medium, and flow conditions are detailed. The proposed microdevice can be used to detect and separate malignant cells from heterogeneous mixture of cells for the purpose of early screening for cancer.     

Capillary electrophoresis with on-chip four-electrode capacitively coupled conductivity detection for application in bioanalysis

Microchip capillary electrophoresis (CE) with integrated four-electrode capacitively coupled conductivity detection is presented. Conductivity detection is a universal detection technique that is relatively independent on the detection pathlength and, especially important for chip-based analysis, is compatible with miniaturization and on-chip integration. The glass microchip structure consists of a 6 cm etched channel (20 microm x 70 microm cross section) with silicon nitride covered walls. In the channel, a 30 nm thick silicon carbide layer covers the electrodes to enable capacitive coupling with the liquid inside the channel as well as to prevent interference of the applied separation field. The detector response was found to be linear over the concentration range from 20 microM up to 2 mM. Detection limits were at the low microM level. Separation of two short peptides with a pI of respectively 5.38 and 4.87 at the 1 mM level demonstrates the applicability for biochemical analysis. At a relatively low separation field strength (50 V/cm) plate numbers in the order of 3500 were achieved. Results obtained with the microdevice compared well with those obtained in a bench scale CE instrument using UV detection under similar conditions.     

Wall-jet conductivity detector for microchip capillary electrophoresis

A new end-column ‘hybrid’ contactless conductivity detector for microchip capillary electrophoresis (CE) was developed. It is based on a “hybrid” arrangement where the receiving electrode is insulated by a thin layer of insulator and placed in the bulk solution of the detection reservoir of the chip, whereas the emitting electrode is in contact with the solution eluted from the channel outlet in a wall-jet arrangement. The favorable features of the new detector including the high sensitivity and low noise, can be attributed to both the direct contact of the ‘emitting’ electrode with the analyte solution as well as to the insulation of the detection electrode from the high DC currents in the electrophoretic circuit. Such arrangement provides a 10-fold sensitivity enhancement compared to currently used on-column contactless conductivity CE microchip detector as well as low values of noise and easy operation. The new design of the wall-jet conductivity detector was tested for separation of explosive-related methylammonium, ammonium, and sodium cations. The new detector design reconsiders the wall-jet arrangement for microchip conductivity detection in scope of improved peak symmetry, simplified study of inter-electrode distance, isolation of the electrodes, position of the wall-jet electrode to the separation channel, baseline stability and low limits of detection.     

Electrowetting on dielectric actuation of droplets with capillary electrophoretic zones for MALDI mass spectrometric analysis

An automated fraction collection interface was developed for coupling CE with MALDI-MS. This fraction collection approach is based on the electrowetting on dielectric (EWOD) phenomenon performed on a digital microfluidic (DMF) board; it does not rely on a MALDI spotter. In this study, a four-peptide mixture was used as a sample test, and the separations were conducted in a portable CE instrument with a 150 μm o.d. × 50 μm i.d. capillary and a contactless conductivity detector. The CE instrument was interfaced with a robust DMF board. The CE fractions were directly deposited onto the DMF board at predetermined locations prior to MALDI analysis. The series of experiments determined the lowest concentration that produces a measurable MALDI signal. The concentrations were 0.25, 0.5, 0.05, and 0.05 nmol for bradykinin, angiotensin, ACTH (18-39), and insulin, respectively. The contactless conductivity detector limit of detection for the same analytes was 2.5 μmol.     

CE frontal analysis employing contactless conductivity detection for determination of CMCs of non-UV absorbing charged surfactants

Micellar systems composed of surfactants are used extensively in academia and industry for many different applications. In this work a highly versatile CE method for determination of CMCs of charged surfactants has been developed. In the case of positively charged surfactants a coating procedure of the fused-silica capillary was used, whereas negatively charged surfactants were analyzed using uncoated capillaries. The CE method is based on frontal analysis (FA) employing use of contactless conductivity and UV detection. The main advantages of the method are that it can be used for non-UV absorbing surfactants without introducing marker compounds which previously has been found to affect CMCs, requires very limited sample volume and is easily implemented and automated using standard CE equipment. The fact that counterions and different aggregated states are separated allows a detailed characterization of the micelle systems using the developed method. In the case of UV absorbing surfactants similar results were obtained employing contactless conductivity and UV detection. Finally, CMCs obtained using conductometry gave similar results as compared to the developed CE-FA procedure.     

Real-time monitoring of strand-displacement DNA amplification by a contactless electrochemical microsystem using interdigitated electrodes

This paper reports the design and implementation of a contactless conductivity detection system which combines a thermal control cell, a data processing system and an electrochemical (EC) cell for label-free isothermal nucleic acid amplification and real-time monitoring. The EC cell consists of a microchamber and interdigitated electrodes as the contactless conductivity biosensor with a cover slip as insulation. In our work, contactless EC measurements, the effects of trehalose on amplification, and chip surface treatment are investigated. With the superior performance of the biosensor, the device can detect the amount of pure DNA at concentrations less than 0.1 pg μl(-1). The EC cell, integrated with a heater and a temperature sensor, has successfully implemented nicking-based strand-displacement amplification at an initial concentration of 2.5 μM and the yields are monitored directly (dismissing the use of probes or labels) on-line. This contactless detector carries important advantages: high anti-interference capability, long detector life, high reusability and low cost. In addition, the small size, low power consumption and portability of the detection cell give the system the potential to be highly integrated for use in field service and point of care applications.     

Rapid determination of partition coefficients of pharmaceuticals by phase distribution and microchip capillary electrophoresis with contactless conductivity detection

A simple microchip CE method integrated with contactless conductivity detection was developed for the direct determination of partition coefficients of selected pharmaceuticals after phase distribution equilibrium. The equilibrium of distribution between two phases for four pharmaceuticals was performed using a 1‐octanol/water system and 1‐octanol/buffer system. During the concentration determination, several major factors affecting detection were investigated in detail for each pharmaceutical to optimize the detection sensitivity. In the optimal conditions, sufficient electrophoretic separation and sensitive detection for each target analyte can be achieved within 40 s. The two systems showed a pH‐dependent partition behavior. Moreover, the measured values showed excellent agreement with those obtained by the traditional shake‐flask method with HPLC–UV detection and literature reports, respectively. The developed method can be successfully applied to measure partition coefficient values of pharmaceuticals and requires much shorter analytical time compared to traditional methods.     

Integration of a contactless conductivity detector into a commercial capillary cassette. Detection of inorganic cations and catecholamines

A contactless conductivity detector integrated into the capillary cassette of Agilent ^3DCE equipment is described. The detector is user-friendly, compact and easily modified. The UV detector of the ^3DCE equipment is available parallel with the contactless conductivity detector increasing the detection power. Two electrolyte solutions, 2-(N-morpholino)ethanesulfonic acid–histidine solution (20 mM, pH 6.0) and ammonium acetate (10 mM, pH 4.0), were used as the separation media for inorganic cations and organic catecholamines, respectively. The detection limit for all metal cations except barium was under 0.5 mg/l, and that for four catecholamines was ca. 10 mg/l. This last value was the same order of magnitude as achieved with parallel UV detection.     

CE frontal analysis based on simultaneous UV and contactless conductivity detection: a general setup for studying noncovalent interactions

CE frontal analysis (CE-FA) has been established as a powerful tool to study noncovalent interactions between macromolecules and small molecules such as drug substances or pharmaceutical excipients. However, when using traditional commercial CE instrumentation, a serious drawback is related to the fact that only UV-active compounds can be studied. In recent years, contactless conductivity detection has become an attractive alternative to UV detection in CE due to its high versatility. In this study, we combine contactless conductivity detection and UV detection in a highly versatile setup for profiling noncovalent interactions between low-molecular-weight molecules and macromolecules. In the case of molecules having a chromophore the setup allows determination of binding constants using two independent detectors. The new contactless conductivity detection cell is compatible with commercial CE instrumentation and is therefore easily implemented in any analysis laboratory with CE expertise.