Elisabeth Verpoorte

Microfluidic chips for clinical and forensic analysis

This review gives an overview of developments in the field of microchip analysis for clinical diagnostic and forensic applications. The approach chosen to review the literature is different from that in most microchip reviews to date, in that the information is presented in terms of analytes tested rather than microchip method. Analyte categories for which examples are presented include (i) drugs (quality control, seizures) and explosives residues, (ii) drugs and endogenous small molecules and ions in biofluids, (iii) proteins and peptides, and (iv) analysis of nucleic acids and oligonucleotides. Few cases of microchip analysis of physiological samples or other “real‐world” matrices were found. However, many of the examples presented have potential application for these samples, especially with ongoing parallel developments involving integration of sample pretreatment onto chips and the use of fluid propulsion mechanisms other than electrokinetic pumping.

Planar microcoil-based microfluidic NMR probes

Microfabricated small-volume NMR probes consisting of electroplated planar microcoils integrated on a glass substrate with etched microfluidic channels are fabricated and tested. 1H NMR spectra are acquired at 300 MHz with three different probes having observed sample volumes of respectively 30, 120, and 470 nL. The achieved sensitivity enables acquisition of an 1H spectrum of 160 microg sucrose in D2O, corresponding to a proof-of-concept for on-chip NMR spectroscopy. Increase of mass-sensitivity with coil diameter reduction is demonstrated experimentally for planar microcoils. Models that enable quantitative prediction of the signal-to-noise ratio and of the influence of microfluidic channel geometry on spectral resolution are presented and successfully compared to the experimental data. The main factor presently limiting sensitivity for high-resolution applications is identified as being probe-induced static magnetic field distortions. Finally, based on the presented model and measured data, future performance of planar microcoil-based microfluidic NMR probes is extrapolated and discussed.

Sample pretreatment on microfabricated devices

The integration of sample pretreatment into microfluidic devices represents one of the remaining hurdles towards achieving true miniaturized total analysis systems (muTAS). The challenge is made more complex by the enormous variation in samples to be analyzed. Moreover, the pretreatment technique has to be compatible with the analysis device to which it is coupled in terms of time, reagent and power consumption, as well as sample volume. This review provides a thorough overview of the developments in this field to date.

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 νm 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 νm and depth of 12 νm. 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 νM) 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.

Comparison of Biocompatibility and Adsorption Properties of Different Plastics for Advanced Microfluidic Cell and Tissue Culture Models

Microfluidic technology is providing new routes toward advanced cell and tissue culture models to better understand human biology and disease. Many advanced devices have been made from poly(dimethylsiloxane) (PDMS) to enable experiments, for example, to study drug metabolism by use of precision-cut liver slices, that are not possible with conventional systems. However, PDMS, a silicone rubber material, is very hydrophobic and tends to exhibit significant adsorption and absorption of hydrophobic drugs and their metabolites. Although glass could be used as an alternative, thermoplastics are better from a cost and fabrication perspective. Thermoplastic polymers (plastics) allow easy surface treatment and are generally transparent and biocompatible. This study focuses on the fabrication of biocompatible microfluidic devices with low adsorption properties from the thermoplastics poly(methyl methacrylate) (PMMA), polystyrene (PS), polycarbonate (PC), and cyclic olefin copolymer (COC) as alternatives for PDMS devices. Thermoplastic surfaces were oxidized using UV-generated ozone or oxygen plasma to reduce adsorption of hydrophobic compounds. Surface hydrophilicity was assessed over 4 weeks by measuring the contact angle of water on the surface. The adsorption of 7-ethoxycoumarin, testosterone, and their metabolites was also determined after UV-ozone treatment. Biocompatibility was assessed by culturing human hepatoma (HepG2) cells on treated surfaces. Comparison of the adsorption properties and biocompatibility of devices in different plastics revealed that only UV-ozone-treated PC and COC devices satisfied both criteria. This paper lays an important foundation that will help researchers make informed decisions with respect to the materials they select for microfluidic cell-based culture experiments.

Sample preconcentration by field amplification stacking for microchip-based capillary electrophoresis.

A microchip structure for field amplification stacking (FAS) was developed, which allowed the formation of comparatively long, volumetrically defined sample plugs with a minimal electrophoretic bias. Up to 20‐fold signal gains were achieved by injection and separation of 400 μm long plugs in a 7.5 cm long channel. We studied fluidic effects arising when solutions with mismatched ionic strengths are electrokinetically handled on microchips. In particular, the generation of pressure‐driven Poiseuille flow effects in the capillary system due to different electroosmotic flow velocities in adjacent solution zones could clearly be observed by video imaging. The formation of a sample plug, stacking of the analyte and subsequent release into the separation column showed that careful control of electric fields in the side channels of the injection element is essential. To further improve the signal gain, a new chip layout was developed for full‐column stacking with subsequent sample matrix removal by polarity switching. The design features a coupled‐column structure with separate stacking and capillary electrophoresis (CE) channels, showing signal enhancements of up to 65‐fold for a 69 mm long stacking channel.

Design and development of a miniaturised total chemical analysis system for on-line lactate and glucose monitoring in biological samples

A miniaturised Total chemical Analysis System (μTAS) for glucose and lactate measurement in biological samples constructed based on an integrated microdialysis sampling and detection system. The complete system incorporates a microdialysis probe for intravascular monitoring in an ex vivo mini-shunt arrangement, and a silicon micromachined stack with incorporated miniaturised flow cell/sensor array. The prototype device has been developed based on state-of-the-art membrane and printed circuit board technology. The flow-through detection system is based on a three-dimensional flow circuit incorporating silicon chips with stacked micromachined channels. An integrated biosensor array (comprising enzyme sensors specific for glucose and lactate) is placed at the base of the stack allowing the detector to be incorporated within the μTAS assembly. These glucose and lactate biosensors are prepared using photolithographic techniques, with measurement based on the detection of hydrogen peroxide at glucose oxidase and lactate oxidase modified platinum electrodes. The resulting amperometric current (at 500 mV vs, Ag/AgCl) is proportional to the concentration of analyte in the sample. All instrumentation is under computer control and the complete unit allows continuous on-line monitoring of glucose and lactate, with fast stable signals over the relevant physiological range for both analytes. The microdialysis system provides 100% sampling efficiency. Sensor performance studies undertaken include optimisation of sensitivity, linearity, operational stability, background current, storage stability and hydration time. The total system (sampling and detection) response time is of the order of 4 min, with sensor sensitivity 1-5 nA mM-1 for glucose and lactate over the range 0.1-33 and 0.05-15 mM, respectively.

Microfluidic Biochip for the Perifusion of Precision-Cut Rat Liver Slices for Metabolism and Toxicology Studies

Early detection of kinetic, metabolic, and toxicity (ADME‐Tox) profiles for new drug candidates is of crucial importance during drug development. This article describes a novel in vitro system for the incubation of precision‐cut liver slices (PCLS) under flow conditions, based on a poly(dimethylsiloxane) (PDMS) device containing 25‐µL microchambers for integration of the slices. The microdevice is coupled to a perifusion system, which enables a constant delivery of nutrients and oxygen and a continuous removal of waste products. Both a highly controlled incubation environment and high metabolite detection sensitivity could be achieved using microfluidics. Liver slices were viable for at least 24 h in the microdevice. The compound, 7‐ethoxycoumarin (7‐EC), was chosen to test metabolism, since its metabolism includes both phase I and phase II metabolism and when tested in the conventional well plate system, correlates well with the in vivo situation (De Kanter et al. 2004. Xenobiotica 34(3): 229–241.). The metabolic rate of 7‐EC was found to be 214 ± 5 pmol/min/mg protein in the microdevice, comparable to well plates, and was constant over time for at least 3 h. This perifusion system better mimics the in vivo situation, and has the potential to significantly contribute to drug metabolism and toxicology studies of novel chemical entities. Biotechnol. Bioeng. 2010;105: 184–194.

Chemical sensing using an integrated microfluidic system based on the Berthelot reaction

Abstract The Berthelot reaction is a well-established colorimetric method for the determination of ammonia. It has been investigated with the particular aim of incorporating it into a simple, reliable analytical microfluidic sensing system. Absorbance measurements for the complex formed when this reaction is performed in microfluidic chips compare very well to those obtained in a spectrophotometric system. The very high reproducibility and efficiency of mixing by diffusion in the microfluidic chip make it a useful tool for future studies of other chemical methods where kinetics are a limiting factor for the response time.

Microfluidic devices for in vitro studies on liver drug metabolism and toxicity

Microfluidic technologies enable the fabrication of advanced in vitro systems incorporating liver tissue or cells to perform metabolism and toxicity studies for drugs and other xenobiotics. The use of microfluidics provides the possibility to utilize a flow of medium, thereby creating a well-controlled microenvironment. The general goals of most in vitro systems in drug research are to optimally mimic the in vivo situation, and to minimize the number of animals required for preclinical studies. Moreover, they may contribute to a reduced attrition rate of drugs at a late stage of the drug development process; this is especially true if human tissue or cells are used. A number of factors are important in achieving good in vivo predictability in microfluidic systems, of which the biological system itself (cells or tissue) and the incubation conditions are the most important. The last couple of years have seen various microfluidic-based in vitro systems being developed to incorporate many different cells and/or tissues. In this review, microfluidics-based in vitro systems realized to study liver metabolism and toxicity are summarized and discussed with respect to their applications, advantages, and limitations. The biological basis of these systems is evaluated, and incubation conditions considered. Precise control of the cell or tissue microenvironment is a key advantage of using microfluidic technologies, and the benefits of exposing the cells to medium flow are demonstrated. Special attention is also paid to the incorporation of multiple cell types or tissues into a microfluidic device for the investigation of interorgan interactions, which are difficult if not impossible to study in conventional systems.