Biomedical subjects

Elisabeth Verpoorte

Publications and source records attributed to Elisabeth Verpoorte.

7 recordsLinked to original sources

PubMed2002

Performance of an integrated microoptical system for fluorescence detection in microfluidic systems.

This article presents a new integrated microfluidic/microoptic device designed for basic biochemical analysis. The microfluidic network is wet-etched in a Borofloat 33 (Pyrex) glass wafer and sealed by means of a second wafer. Unlike other similar microfluidic systems, elements of the detection system are realized with the help of microfabrication techniques and directly deposited on both sides of the microchemical chip. The detection system is composed of the combination of refractive circular or elliptical microlens arrays and chromium aperture arrays. The microfluidic channels are 60 microm wide and 25 microm deep. The elliptical microlenses have a major axis of 400 microm and a minor axis of 350 microm. The circular microlens diameters range from 280 microm to 350 microm. The apertures deposited on the outer chip surfaces are etched in a 3000-A-thick chromium layer. The overall thickness of this microchemical system is < 1.6 mm. A limit of detection of 3.3 nM for a Cy5 solution in phosphate buffer (pH 7.4) was demonstrated. The cross-talk signal measured between two adjacent microchannels with 1 mm pitch was < 1:5600, meaning that < or = 1.8 x 10(-4)% of the fluorescence light power emitted from one microchannel filled with a 50 microM Cy5 solution reaches the photodetector at the adjacent microchannel. This performance compares very well with that obtainable in microchemical chips using confocal fluorescence systems, taking differences in parameters, such as excitation power into microchannels, data acquisition rates, and signal filtering into account.

Journal Article↗

PubMed2002

Analysis of lipoproteins by capillary zone electrophoresis in microfluidic devices: assay development and surface roughness measurements.

The development of a new assay for lipoproteins by capillary electrophoresis in fused-silica capillaries and in glass microdevices is described in this paper. The separation of low-density (LDL) and high-density (HDL) lipoproteins by capillary zone electrophoresis is demonstrated in fused-silica capillaries with both UV absorption and laser-induced fluorescence detection. This separation was accomplished using Tricine buffer (pH 9.0) with methylglucamine added as a dynamic coating. With UV detection, LDL eluted as a relatively sharp peak with a migration time of approximately 11 min and HDL eluted as a broad peak with a migration time of 12.5 min. Fluorescence detection of lipoproteins stained with NBD-ceramide was used with the same buffer system to give comparable results. Furthermore, fluorescence staining of human serum samples yielded results similar to the fluorescently stained LDL and HDL fractions, showing that this method can be used to quantify lipoproteins in serum samples. The method was also used to detect lipoproteins in glass micro-CE devices. Very similar results were obtained in microdevices although with much faster analysis times, LDL eluted as a sharp peak at approximately 25 s and HDL as a broad peak at slightly longer time. In addition, higher resolution was obtained on chips. To our knowledge, these results show the first separation and detection of lipoproteins in a microfluidic device using native serum samples. Atomic force microscopy was used to characterize the rms surface roughness (Rq) of microfluidic channels directly. Devices with different surface roughness values were fabricated using two different etchants for Pyrex wafers with a polysilicon masking layer. Using 49% HF, the measured roughness is Rq = 10.9 +/- 1.6 nm and with buffered HF (NH4F + HF) the roughness is Rq = 2.4 +/- 0.7 nm. At this level of surface roughness, there is no observable effect on the performance of the devices for this lipoprotein separation.

Chromatography, Gel↗

PubMed2002

An integrated fritless column for on-chip capillary electrochromatography with conventional stationary phases.

A new polymer device for use with conventional particulate stationary phases for on-chip, fritless, capillary electrochromatography (CEC) has been realized. The structure includes an injector and a tapered column in which the particles of the stationary phase are retained and stabilized. The chips were easily fabricated in poly(dimethylsiloxane) using deep-reactive-ion-etched silicon masters, and tested using a capillary electrophoretic separation of FITC-labeled amino acids. To perform CEC, the separation channel was packed using a vacuum with 3-microm, octadecylsilanized silica microspheres. The packing was stabilized in the column by a thermal treatment, and its stability and quality were evaluated using in-column indirect fluorescence detection. The effects of voltage on electro-osmotic flow and on efficiency were investigated, and the separation of two neutral compounds was achieved in less than 15 s.

Amino Acids↗

PubMed2002

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.

Antibodies↗

PubMed2002

Application of surface biopassivated disposable poly(dimethylsiloxane)/glass chips to a heterogeneous competitive human serum immunoglobulin G immunoassay with incorporated internal standard.

A microfluidic platform for a heterogeneous competitive immunoassay of human immunoglobulin G (IgG) employing Cy5-human IgG as tracer and Cy3-mouse IgG as internal standard was developed. The device consisted of microchannels made of poly(dimethylsiloxane) and glass which were patterned with antibodies against human IgG and mouse IgG. Electrokinetic sample transport was employed in order to exploit the small difference between the net mobilities of analyte and tracer, thereby achieving favorable conditions for the performance of the competitive immunoreaction. The overall quality of the disposable chip and performance of the immunoassay were controlled by monitoring the fluorescence of bound tracer and bound internal standard. Analyses with an insufficient internal standard response were discarded, and immunoassay data evaluation was based on the ratio of tracer and internal standard fluorescence. Using synthetic samples in the range from 0 to 80 microg/mL IgG and alkaline running conditions, a concentration-dependent response with reproducible Cy5/Cy3 signal ratios (average relative standard deviation of 6.8%) was obtained. Chips stored with solution in the channels at 4 degrees C over a two-month period were found to perform like freshly prepared chips, whereas chips stored dry at -20 degrees C and rehydrated prior to use could not be employed. The analysis of patient sera showed that the immunoassay platform behaved differently in the presence of serum-based samples. Using the same conditions as for the synthetic samples, no concentration dependence was noted. With a large excess of tracer, however, an IgG concentration dependence was observed, permitting distinction of samples of patients with normal IgG serum levels (8-16 mg/mL) from those with elevated IgG concentrations (>16 mg/mL).

Animals↗

PubMed2002

Low-density lipoprotein analysis in microchip capillary electrophoresis systems.

Due to the mounting evidence for altered lipoprotein and cholesterol-lipoprotein content in several disease states, there has been an increasing interest in analytical methods for lipoprotein profiling for diagnosis. The separation of low- and high-density lipoproteins (LDL and HDL, respectively) has been recently demonstrated using a microchip capillary electrophoresis (CE) system [1]. In contrast to this previous study, the present report demonstrates that LDL analysis can be performed in an uncoated glass microchannel. Moreover, by adding sodium dodecyl sulfate (SDS) to the sample at a concentration well below the critical micellar concentration prior to injection, the LDL peak undergoes a focusing effect and exhibits an apparent efficiency of 2.2 x 10(7) plates/m. Laser light scattering experiments demonstrate that the low concentration of SDS used does not significantly alter lipoprotein particle size distribution within the time course that the analysis is performed. It is thus hypothesized that SDS nondisruptively coats LDL particles. The peak sharpening effect, observed only when SDS is added solely to the sample, is probably due to a mobility gradient created between the sample and the running buffer. The chip-based method demonstrated here has the potential for rapid analysis and sensitive detection of different LDL forms of clinical relevance.

Electrophoresis, Capillary↗

PubMed2002

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.

Cations↗