Microchip Electrophoresis Devices Research Articles

A fully functional palmtop microchip electrophoresis analyzer with laser-induced fluorescence detection

Microchip electrophoresis (MCE) is an ideal tool for rapid on-site analysis of multiple components, therefore the demand for portable MCE devices is increasing. In this work, a fully functional palmtop MCE analyzer was built, integrating units of sample injection, laser-induced fluorescence (LIF) detection, high-voltage (HV), imaging and data acquisition. Frames were fabricated with 3D printing to encapsulate miniaturized components selected with balanced consideration on size, cost and performance. The LIF unit contains a laser diode as the excitation light source, a collimating lens used in the commercial laser diode as the focusing lens and an avalanche photodiode (APD) for the laser distance meter as the photoelectric converter. The sample injection was achieved by negative pressure induced pinching. The total cost of the device is approximately $600 with a size of 89 × 98 × 98 mm3 and a weight of 350 g. It can be powered by a laptop or a mobile power bank. The limit of detection (LOD) measured under a typical MCE condition for sodium fluorescein could be 110 pM. The applicability of the device was demonstrated with the detection of rhodamines and DNA fragments.

Electrokinetic characterization of hybrid NOA 81-glass microchips: Application to protein microchip electrophoresis with indirect fluorescence detection.

A low-cost and straightforward hybrid NOA (Norland optical adhesive) 81-glass microchip electrophoresis device was designed and developed for protein separation using indirect fluorescence detection. This new microchip was first characterized in terms of surface charge density via electroosmotic mobility measurement and stability over time. A systematic determination of the electroosmotic mobility (μeo ) over a wide pH range (2-10) and at various ionic strengths (20-50mM) was developed for the first time via the neutral marker approach in an original simple frontal methodology. The evolution of μeo was proved consistent with the silanol and thiol functions arising from the glass and the NOA materials, respectively. The repeatability and reproducibility of the measurements on different microchips (RSD<14%) and within 15 days (less than 5% decrease) were successfully demonstrated. The microchip was then applied for the efficient electrophoretic separation of proteins in a zonal mode coupled with indirect fluorescence detection, which is, to our knowledge, the first proof of concept of capillary zone electrophoresis in this hybrid microsystem.

3D-printed microchip electrophoresis device containing spiral electrodes for integrated capacitively coupled contactless conductivity detection.

In this work, we demonstrate for the first time the design and fabrication of microchip electrophoresis devices containing cross-shaped channels and spiral electrodes around the separation channel for microchip electrophoresis and capacitively coupled contactless conductivity detection. The whole device was prepared in a digital light processing-based 3D printer in poly(ethylene glycol) diacrylate resin. Outstanding X-Y resolution of the customized 3D printer ensured the fabrication of 40-μm cross section channels. The spiral channels were filled with melted gallium to form conductive electrodes around the separation channel. We demonstrate the applicability of the device on the separation of sodium, potassium, and lithium cations by microchip electrophoresis. Graphical abstract.

Separation of carbohydrates on electrophoresis microchips with controlled electrolysis.

This study reports the separation of fructose, galactose, glucose, lactose and sucrose on glass microchip electrophoresis (ME) devices using a microfluidic platform adapted with external reservoirs for controlling the electrolysis phenomenon. The connections between external reservoirs and microfluidic platform were performed by saline bridges created using silicone tubing filled with BGE. The separation conditions were optimized and the best results were achieved using a BGE containing 75 mmol/L NaOH and 15 mmol/L trisodium phosphate. Electrophoretic separations were monitored using a capacitively coupled contactless conductivity detection system. The controlled electrolysis has successfully allowed the application of a higher voltage on the separation channel promoting the baseline separation of five carbohydrates within 180 s with great run-to-run repeatability (RSD < 1%). The achieved efficiencies ranged from 45 000 ±6000 to 70 000 ±3000 plates/m demonstrating a performance better than ME devices without controlled electrolysis. The proposed system offered good linearity from 1 to 10 mmol/L and LODs between 150 and 740 μmol/L. The use of external tubes for controlling the electrolysis phenomenon on ME devices has solved common problems associated to run-to-run repeatability and analytical reliability required for routine and quantitative analysis.

Sample Injection Techniques.

Sample introduction is an important consideration in any microchip electrophoresis (ME) separation. The number of applications and level of complexity of ME devices continue to increase, but the introduction of sample into the separation channel has remained relatively constant. This work describes the most common electrophoretic methods of performing sample injection for ME applications using electroosmotic flow (EOF), providing both a transition from capillary electrophoresis (CE) separations and a straightforward entry point into ME.

Paper-Based Electrophoresis Microchip as a Powerful Tool for Bioanalytical Applications.

This chapter describes the development of paper-based microchip electrophoresis (pME) devices for the separation of clinically relevant compounds. pME were fabricated by laser cut and thermal lamination process using polyester pouches. In addition, hand-drawn pencil electrodes were integrated to the device to perform capacitively coupled contactless conductivity detection (C4D). Finished device costs less than US$ 0.10 and did not require either sophisticated instrumentation or clean room facilities. Furthermore, pME is lightweight, easy to handle, flexible, and robust. pME-C4D device revealed an excellent capacity to separate BSA and creatinine in less than 150s with baseline resolution. The device proposed in this chapter has proven to be a good alternative as a platform for the diagnosis of diseases from renal disorders such as diabetes mellitus and heart disease.

Rapid separation of post‐blast explosive residues on glass electrophoresis microchips

This study describes the development of an analytical methodology based on the use of microchip electrophoresis (ME) devices integrated with capacitively coupled contactless conductivity detection (C4 D) for the separation and detection of inorganic anions in post-blast explosive residues. The best separation condition was achieved using a running buffer composed of 35mmol/L lactic acid, 10mmol/L histidine and 0.070mmol/L cetyl(trimethyl ammonium) bromide. For C4 D measurements, the highest sensitivity was obtained applying a 700kHz sinusoidal wave with excitation voltage of 20Vpp . The separation of Cl- , NO3 - , NO2 - , SO4 2- , ClO4 - and ClO3 - was performed within ca. 150 s with baseline resolution and efficiencies between 4.4 × 104 and 1.7 × 105 plates/m. The found limits of detection ranged between 2.5 and 9.5μmol/L. Last, real samples of post-blast explosive residues were analyzed on the ME-C4 D devices obtaining successfully the determination of Cl- , NO3 - and SO4 2- . The achieved concentration values varied between 12.8-72.5mg/L for Cl- , 1.7-293.1mg/L for NO3 - and 1.3-201.3 mg/L for SO4 2- . The data obtained using ME-C4 D devices were in good agreement with the concentrations found by ion chromatography. The approach reported herein has provided short analysis time, instrumental simplicity, good analytical performance and low cost. Furthermore, the ME-C4 D devices emerge as a powerful and portable analytical platform for on-site analysis demonstrating to be a promising tool for the crime scene investigation.

A Microchip Electrophoresis Device Integrated with the Top-bottom Antiparallel Electrodes of Indium Tin Oxide to Detect Inorganic Ions by Contact Conductivity.

A microchip electrophoresis (ME) device with off-channel contact conductivity detection (C2D) was constructed using top-bottom antiparallel indium tin oxide (ITO) electrodes and a cross-type microchannel. The 500-m wide top-bottom antiparallel decouplers were found to effectively decrease the interference of the electrophoretic current. The cross-type microchannel was formed by bonding the patterned negative photoresist microstructures and the two top-bottom opposed ITO-deposited glass substrates. Five seconds of 150 V/cm injection field and the 100 V/cm separation field equipped with the C2D of AC 200 mV excitation voltage provided adequate ME operational parameters to obtain the K+ and Na+ peaks separation. The ME devices obtained good coefficients in a range of 11000 μM for the K+ and Na+ detection. The calculated limit of detection was 1 μM. This design for off-channel and top-bottom antiparallel electrodes shows that C2D ME devices have great potential for the measurement of inorganic ions.

Screening of seized cocaine samples using electrophoresis microchips with integrated contactless conductivity detection.

This study describes the development of a new analytical method for the separation and detection of cocaine (COC) and its adulterants, or cutting agents, using microchip electrophoresis (ME) devices coupled with capacitively coupled contactless conductivity detection (C4 D). All the experiments were carried out using a glass commercial ME device containing two pairs of integrated sensing electrodes. The running buffer composed of 20 mmol/L amino-2-(hydroxymethyl) propane-1,3-diol and 10 mmol/L 3,4-dimethoxycinnamic acid provided the best separation conditions for COC and its adulterants with baseline resolution (R>1.6), separation efficiencies ranging from (2.9 ± 0.1) to (3.2 ± 0.2) × 105 plates/m, and estimated LOD values between 40 and 150 μmol/L. The quantification of COC was successfully performed in four samples seized by the Brazilian Federal Police Department and all predicted values agree with values estimated by the reference method. Some other interfering species were detected in the seized samples during the screening procedure on ME-C4 D devices. While lidocaine was detected in sample 3, the presence of levamisole was observed in samples 2 and 4. However, their concentrations were estimated to be below the LOQ. ME-C4 D devices have proved to be quite efficient for the identification and quantification of COC with errors lower than 10% when compared to the data obtained by a reference method. The approach herein reported offers great potential to be used for on-site COC screening in seized samples.

Toward miniaturized analysis of chemical identity and purity of radiopharmaceuticals via microchip electrophoresis.

Miniaturized synthesis of positron emission tomography (PET) tracers is poised to offer numerous advantages including reduced tracer production costs and increased availability of diverse tracers. While many steps of the tracer production process have been miniaturized, there has been relatively little development of microscale systems for the quality control (QC) testing process that is required by regulatory agencies to ensure purity, identity, and biological safety of the radiotracer before use in human subjects. Every batch must be tested, and in contrast with ordinary pharmaceuticals, the whole set of tests of radiopharmaceuticals must be completed within a short-period of time to minimize losses due to radioactive decay. By replacing conventional techniques with microscale analytical ones, it may be possible to significantly reduce instrument cost, conserve lab space, shorten analysis times, and streamline this aspect of PET tracer production. We focus in this work on miniaturizing the subset of QC tests for chemical identity and purity. These tests generally require high-resolution chromatographic separation prior to detection to enable the approach to be applied to many different tracers (and their impurities), and have not yet, to the best of our knowledge, been tackled in microfluidic systems. Toward this end, we previously explored the feasibility of using the technique of capillary electrophoresis (CE) as a replacement for the "gold standard" approach of using high-performance liquid chromatography (HPLC) since CE offers similar separating power, flexibility, and sensitivity, but can readily be implemented in a microchip format. Using a conventional CE system, we previously demonstrated the successful separation of non-radioactive version of a clinical PET tracer, 3'-deoxy-3'-fluorothymidine (FLT), from its known by-products, and the separation of the PET tracer 1-(2'-deoxy-2'-fluoro-β-D-arabinofuranosyl)-cytosine (D-FAC) from its α-isomer, with sensitivity nearly as good as HPLC. Building on this feasibility study, in this paper, we describe the first effort to miniaturize the chemical identity and purity tests by using microchip electrophoresis (MCE). The fully automated proof-of-concept system comprises a chip for sample injection, a separation capillary, and an optical detection chip. Using the same model compound (FLT and its known by-products), we demonstrate that samples can be injected, separated, and detected, and show the potential to match the performance of HPLC. Addition of a radiation detector in the future would enable analysis of radiochemical identity and purity in the same device. We envision that eventually this MCE method could be combined with other miniaturized QC tests into a compact integrated system for automated routine QC testing of radiopharmaceuticals in the future. Graphical abstract Miniaturized quality control (QC) testing of batches of radiopharmaceuticals via microfluidic analysis. The proof-of-concept hybrid microchip electrophoresis (MCE) device demonstrated the feasibility of achieving comparable performance to conventional analytical instruments (HPLC or CE) for chemical purity testing.

Sensitivity enhancing injection from a sample reservoir and channel interface in microchip electrophoresis.

The stacking of a cationic analyte (i.e., rhodamine B) at the interface between a sample reservoir and channel in a microchip electrophoresis device is described for the first time. Stacking at negative polarity was by micelle to solvent stacking where the dye was prepared in a micellar solution (5 mM sodium dodecyl sulfate in 25 mM phosphoric acid, pH 2.5) and the channel was filled with high methanol content background solution (70% methanol in 50 mM phosphoric acid, pH 2.5). The injection of the stacked dye into the channel was by simple reversal of the voltage polarity with the sample solution and background solution at the anodic and cathodic reservoirs of the straight channel, respectively. The enrichment of rhodamine B at the interface and injection of the stacked dye into the channel was clearly visualized using an inverted fluorescence microscope. A notable sensitivity enhancement factor of up to 150 was achieved after 2 min at 1 kV of micelle to solvent stacking. The proposed technique will be useful as a concentration step for analyte mixtures in simple and classical cross-channel microchip electrophoresis devices or for the controlled delivery of enriched reagents or analytes as narrow plugs in advanced microchip electrophoresis devices.

Simple approach for the fabrication of screen-printed carbon-based electrode for amperometric detection on microchip electrophoresis

This paper describes a simple method for the fabrication of screen-printed based electrodes for amperometric detection on microchip electrophoresis (ME) devices. The procedure developed is quite simple and does not require expensive instrumentation or sophisticated protocols commonly employed on the production of amperometric sensors, such as photolithography or sputtering steps. The electrodes were fabricated through manual deposition of home-made conductive carbon ink over patterned acrylic substrate. Morphological structure and electrochemical behavior of the carbon electrodes were investigated by scanning electron microscopy and cyclic voltammetry. The produced amperometric sensors were coupled to polydimethylsiloxane (PDMS) microchips at end-channel configuration in order to evaluate their analytical performance. For this purpose, electrophoretic experiments were carried out using nitrite and ascorbic acid as model analytes. Separation of these substances was successfully performed within 50s with good resolution (R = 1.2) and sensitivities (713.5 pA/μM for nitrite and 255.4 pA/μM for ascorbate). The reproducibility of the fabrication method was evaluated and revealed good values concerning the peak currents obtained (8.7% for nitrite and 9.3% for ascorbate). The electrodes obtained through this method exhibited satisfactory lifetime (ca. 400 runs) over low fabrication cost (less than $1 per piece). The feasibility of the proposed device for real analysis was demonstrated through the determination of nitrite concentration levels in drinking water samples. Based on the results achieved, the approach proposed here shows itself as an interesting alternative for simple fabrication of carbon-based electrodes. Furthermore, the devices indicate great promise for other kind of analytical applications involving ME devices.

Determination of anti-oxidative histidine dipeptides in poultry by microchip capillary electrophoresis with contactless conductivity detection.

A home-made microchip electrophoresis (MCE) device was used to quantitate two biologically important histidine dipeptides, carnosine and anserine, using capacitively coupled contactless conductivity detection (C4D), at pH 2.7. The C4D detector exhibited a linear response to both carnosine and anserine in the range of 0-200μM for the individual dipeptides and in the range of 0-100μM for each dipeptide when both were present as a mixture. The limit of detections (LOD) for the dipeptides in the mixture were 0.10μM for carnosine and 0.16μM for anserine. Standard addition was used to detemine the accuracy of the method. For carnosine and anserine the recoveries were in the range of 96.7±4.9-106.0±7.5% and 95.3±4.5-105.0±5.1% in thigh muscle and 97.5±5.1-105.0±7.5% and 95.3±5.4-97.3±5.6% in breast muscle, respectively.

Authenticity screening of seized whiskey samples using electrophoresis microchips coupled with contactless conductivity detection.

This report describes for the first time the use of microchip electrophoresis (ME) devices integrated with capacitively coupled contactless conductivity detection (C4 D) to investigate the authenticity of seized whiskey samples, which were probably adulterated by simple dilution with tap water. The proposed microfluidic platform was explored for the monitoring of anionic species (Cl- and F- ) in both original and tampered samples. The best separations were achieved within 70 s using a running buffer composed of lactic acid and histidine (pH = 5.9). ME-C4 D devices were used to analyze samples from three different brands (five samples each). Based on the presence of inorganic anions like Cl- , F- , SO42- and NO2- in different amounts, the authenticity of seized whiskeys was compared to original samples. According to the reported data, the proposed microfluidic platform can be useful to help regulatory authorities in the investigation and monitoring of authenticity of commercialized whiskey beverages.

Hydrodynamic injection on electrophoresis microchips using an electronic micropipette

Here we report for the first time the use of an electronic micropipette as hydrodynamic (HD) injector for microchip electrophoresis (ME) devices. The micropipette was directly coupled to a PDMS device, which had been fabricated in a simple cross format with two auxiliary channels for sample volume splitting. Sample flow during the injection procedure was controlled in automatic dispenser mode using a volume of 0.6µL. Channel width and device configuration were optimized and the best results were achieved using a simple cross layout containing two auxiliary channels with 300µm width for sample splitting. The performance of the HD injector was evaluated using a model mixture of high-mobility cationic species. The results obtained were compared to the data obtained via electrokinetic (EK) injection. Overall, the HD provided better analytical performance in terms of resolution and injection-to-injection repeatability. The relative standard deviation (RSD) values for peak intensities were lower than 5% (n=10) when the micropipette was employed. In comparison with EK injection, the use of the proposed HD injector revealed an unbiased profile for a mixture containing K+ and Li+(300 µmol L−1 each) over various buffer concentrations. For EK injection, the peak areas decreased from 2.92 ± 0.20–0.72 ± 0.14Vs for K+ and from 1.30 ± 0.10–0.38 ± 0.10Vs for Li+ when the running buffer increased from 20 to 50mmolL−1. For HD injection, the peak areas for K+ and Li+ exhibited average values of 2.48±0.07 and 2.10±0.06Vs, respectively. The limits of detection (LDs) for K+, Na+ and Li+ ranged from 18 to 23µmolL−1. HD injection through an electronic micropipette allows to automatically dispense a bias-free amount of sample inside microchannels with acceptable repeatability. The proposed approach also exhibited instrumental simplicity, portability and minimal microfabrication requirements.

Contactless conductivity detection for analytical techniques- Developments from 2014 to 2016.

The development of capacitively coupled contactless conductivity detection for the two-year period from mid-2014 to mid-2016 is covered in this review. This includes a survey of fundamental studies and further developments of the measuring technique reported as well as a discussion of new applications. These mostly concern capillary electrophoresis carried out in conventional capillaries as well as on microchip electrophoresis devices. The main focus is on the determination of small non-UV-absorbing organic ions and inorganic ions in different types of samples of clinical, nutritional or environmental interest. Outside of electrophoresis contactless conductivity detection is finding uses in detection in column chromatography, flow-injection analysis and industrial applications.

Monitoring of nitrite, nitrate, chloride and sulfate in environmental samples using electrophoresis microchips coupled with contactless conductivity detection

This report describes the development of an analytical methodology on microchip electrophoresis (ME) devices coupled with capacitively coupled contactless conductivity detection (C4D) to monitor inorganic anions in environmental samples. The buffer composition as well as detection operating parameters were optimized to achieve the best separation selectivity and detector sensitivity, respectively. Electrophoretic separations of Cl−, NO3−, SO42− and NO2− were successfully performed within 60s using a running buffer composed of 30mmolL−1 latic acid and 15mmolL−1l-histidine (His). The best detectability levels were found applying a sinusoidal wave with 1100-kHz-frequency and 60-Vpp amplitude. Quantitative analyzes of inorganic anions were carried out in the presence of Cr2O72− ion as internal standard (IS), which ensured great repeatability in terms of migration times (<1%) and peak areas (6.2–7.6%) for thirty consecutive injections. The analytical performance revealed a linear behavior for concentration ranges between 0–120μmolL−1 (Cl−, NO2− and NO3−) and 0–60μmolL−1 (SO42−) and limits of detection (LODs) varying from 2.0 to 4.9μmolL−1. The concentration levels of anionic species were determined in aquarium, river and biofertilizer samples with recovery values between 91% and 105%. The nitrification steps associated with conversion of ammonium to nitrite followed by the conversion of nitrite to nitrate were successfully monitored in a simulated environment without fishes during a period of twelve weeks. Lastly, the monitoring of anionic species was carried out during eight weeks in an aquarium environment containing ten fishes from Danio rerio (Ciprynidae). The recorded data revealed the absence of nitrite and a gradual increase on the ammonium and nitrate concentration levels during eight weeks, thus suggesting the direct conversion of ammonium to nitrate. Based on the data herein reported, the proposed analytical methodology can be used for routine environmental analysis.

Electrodeposition of reduced graphene oxide on a Pt electrode and its use as amperometric sensor in microchip electrophoresis.

This report describes the development and application of a novel graphene-modified electrode to be used as amperometric sensor in microchip electrophoresis (ME) devices. The modified electrode was achieved based on electroreduction of graphene oxide on an integrated Pt working electrode of a commercial ME device. The surface modification was characterized by SEM and cyclic voltammetry techniques. The results indicated that graphene sheets were successfully deposited exhibiting higher surface conductivity and greater electrode sensitivity. The performance of the modified electrode for the amperometric detection on ME devices has been demonstrated by the separation and detection of an anionic mixture containing iodide and ascorbate. The graphene-modified electrode provided significantly higher sensitivity (896.7 vs. 210.9 pA/μM for iodide and 217.8 vs. 127.8 pA/μM for ascorbate), better separation efficiencies (3400 vs. 700 plates/m for iodide and 10000 vs. 2400 plates/m for ascorbate), enhanced peak resolutions (1.6 vs. 1.0), and LODs (1.5 vs. 5.3 μM for iodide and 3.1 vs. 7.3 μM for ascorbate) in comparison with the unmodified Pt electrode. The proposed amperometric sensor was successfully applied for the analysis of ascorbic acid (through its anionic form) in a commercial medicine sample, and the results achieved were in agreement with the value provided by the supplier. Based on the data here presented, the modified graphene electrode shows great promise for ME applications.

Evaluation of in-channel amperometric detection using a dual-channel microchip electrophoresis device and a two-electrode potentiostat for reverse polarity separations: Microfluidics and Miniaturization

Evaluation of in-channel amperometric detection using a dual-channel microchip electrophoresis device and a two-electrode potentiostat for reverse polarity separations: Microfluidics and Miniaturization

Evaluation of in-channel amperometric detection using a dual-channel microchip electrophoresis device and a two-electrode potentiostat for reverse polarity separations.

In-channel amperometric detection combined with dual-channel microchip electrophoresis is evaluated using a two-electrode isolated potentiostat for reverse polarity separations. The device consists of two separate channels with the working and reference electrodes placed at identical positions relative to the end of the channel, enabling noise subtraction. In previous reports of this configuration, normal polarity and a three-electrode detection system were used. In the two-electrode detection system described here, the electrode in the reference channel acts as both the counter and reference. The effect of electrode placement in the channels on noise and detector response was investigated using nitrite, tyrosine, and hydrogen peroxide as model compounds. The effects of electrode material and size and type of reference electrode on noise and the potential shift of hydrodynamic voltammograms for the model compounds were determined. In addition, the performance of two- and three-electrode configurations using Pt and Ag/AgCl reference electrodes was compared. Although the signal was attenuated with the Pt reference, the noise was also significantly reduced. It was found that lower LOD were obtained for all three compounds with the dual-channel configuration compared to single-channel, in-channel detection. The dual-channel method was then used for the detection of nitrite in a dermal microdialysis sample obtained from a sheep following nitroglycerin administration.