Biomedical subjects

Jacques Josserand

Publications and source records attributed to Jacques Josserand.

12 recordsLinked to original sources

PubMed2005

Dynamic protein adsorption in microchannels by "stop-flow" and continuous flow.

This work addresses two ways of loading proteins on microchannel surfaces for immunoassay applications: the "stop-flow" and the continuous flow processes. The "stop-flow" method consists of successive static incubation periods where the bulk solution depletes upon the adsorption process. In the present paper, a multi-step "stop-flow" protein coating is studied and compared to a coating under continuous flow conditions. For the "stop-flow", a non-dimensional parameter is here introduced, indicating the adsorbing capacity of the system, by which it is possible to calculate the number of loads necessary to reach the optimum coverage. For the continuous flow, the effects on the adsorption of the kinetic rates, flow velocity and wall capacity have been considered. This study shows the importance of a careful choice of the fluid velocity to minimise the sample waste. For diffusion controlled and kinetics controlled processes, two flow velocity criteria are provided in order to obtain the best possible coverage, with the same amount of sample as with the "stop-flow".

Adsorption↗

PubMed2005

Numerical simulation of two-phase partition chromatography in microchannels for moderated log P measurements.

A finite element simulation has been used in order to study the partition chromatography process of one species between an aqueous mobile phase and an organic stationary phase located at the bottom of a rectangular microchannel. The transient model incorporates convection--diffusion of the species in the water phase coupled to the diffusion in the stationary organic phase by the way of the partition kinetics at the interface. The time evolution of the injected species concentration is analyzed versus the velocity of the mobile phase, the detecting position and the thickness of the stationary phase. The comparison of simulation results with both experimental data and analytical model confirm its validity. These simulations show that thin channels can be used to measure log P of molecules from their retention time. Finally, we have shown how the sample velocity can be optimized for a given geometry of the channel and diffusion coefficient of the species.

Calibration↗

PubMed2005

Salt removal during Off-Gel electrophoresis of protein samples.

The Off-Gel technology was recently described for protein fractionation in a solution placed on top of an immobilized pH gradient gel. In addition, this process was found to remove salts from the biological samples to analyze. This desalting effect is studied experimentally in a conductometric prototype cell. A simplified analytical model is developed to understand this process and a good agreement is found with the conductivity measurements. To illustrate the desalting of a biological sample, a 1 mg.mL(-1) solution of beta-lactoglobulin A in 0.1 M NaCl is subjected to electrophoresis in a single compartment Off-Gel cell. The analysis of the resulting sample by ESI-MS demonstrates the effective removal of salt. A finite element diffusion-migration model is also used to illustrate how the nonuniformity of the electric field in the cell, associated with the salt migration, can slow down the desalting process.

Electric Conductivity↗

PubMed2004

Protein adsorption in static microsystems: effect of the surface to volume ratio.

A numerical model for the adsorption kinetics of proteins on the walls of a microchannel has been developed using the finite element method (FEM) to address the coupling with diffusion phenomena in the restricted microchannel volume. Time evolutions of the concentration of one species are given, both in solution and on the microchannel walls. The model illustrates the adsorption limitation sometimes observed when the microdimensions of these systems induce a global depletion of the bulk solution. A new non-dimensional parameter is introduced to predict the final value of the coverage of any microsystem under static adsorption. A working curve and a criteria (h/K[Gamma](max) > 10) are provided in order to choose, for given adsorption characteristics, the value of the volume-to-surface ratio (i.e. the channel height h) avoiding depletion effects on the coverage (relative coverage greater than 90% of the theoretical one). Simulations were compared with confocal microscopy measurements of IgG antibody adsorption on the walls of a PET microchannel. The fit of the model to the experimental data show that the adsorption is under apparent kinetic control.

Adsorption↗

PubMed2004

Passive conductivity detection for capillary electrophoresis.

A passive electrochemical detection principle that can be applied to capillary electrophoresis is presented. The separation electrical field is used to generate a potential difference between two electrodes located along the channel. For constant-current electrophoresis, the generated signal is proportional to the resistance of the solution passing between the two electrodes. Contrary to conductivity detectors that are ac driven and need to be decoupled from the separation field, the passive detection directly takes advantage of the separation field. The signal is simply measured by a high-impedance voltmeter. The detection concept has been validated by numerical simulations showing how the magnitude of the signal is related to the ratio between the electrode distance and the length of the sample plug. As a proof of the principle, this detection concept has been demonstrated by the electrophoretic separation of three alkali ions on a polymer microchip. Based on preliminary results, a detection limit of 20 microM and a dynamic range of up to 3 orders of magnitude have been achieved.

Journal Article↗

PubMed2004

Generation of mass tags by the inherent electrochemistry of electrospray for protein mass spectrometry.

We present herein a review of our work on the on-line electrochemical generation of mass tags toward cysteine residues in peptides and proteins. Taking advantage of the inherent electrochemical nature of electrospray generated from a microfabricated microspray emitter, selective probes for cysteine were developed and tested for on-line nonquantitative mass tagging of peptides and proteins. The nonquantitative aspect of the covalent tagging thus allows direct counting of free cysteines in the mass spectrum of a biomolecule through additional adduct peaks. Several substituted hydroquinones were investigated in terms of electrochemical properties, and their usefulness for on-line mass tagging during microspray experiments were assessed with L-cysteine, peptides, and intact proteins. Complementarily, numerical simulations were performed to properly understand the respective roles of mass transport, kinetics of electrochemical-chemical reactions, and design of the microspray emitter in the mass tagging overall efficiency. Finally, the on-line electrochemical tagging of cysteine residues was applied to the analysis of tryptic peptides of purified model proteins for protein identification through peptide mass fingerprinting.

Cysteine↗

PubMed2003

Numerical investigation of an electrochemically induced tagging in a nanospray for protein analysis.

An on-line tagging of target species is simulated using the finite element method. A numerical model of an electrochemical EC(2X)E mechanism in a flow channel cell has been developed, corresponding to the electrochemical generation of a probe and the subsequent homogeneous reaction with the target. The kinetic and convective aspects are validated on short electrode geometries before taking into account the depletion of the target species. The model is then assessed according to previous experimental results on the on-line tagging of proteins. The occurrence and the efficiency of the on-line tagging are studied for both pressure-driven and electroosmotic flows. The involved phenomena including kinetic aspects are described in detail. Finally, optimal conditions for an effective quantitative tagging are discussed.

Algorithms↗

PubMed2003

Microfluidic systems in proteomics.

We present the state-of-the-art in miniaturized sample preparation, immunoassays, one-dimensional and multidimensional analyte separations, and coupling of microdevices with electrospray ionization-mass spectrometry. Hyphenation of these different techniques and their relevance to proteomics will be discussed. In particular, we will show that analytical performances of microfluidic analytical systems are already close to fulfill the requirements for proteomics, and that miniaturization results at the same time in a dramatic increase in analysis throughput. Throughout this review, some examples of analytical operations that cannot be achieved without microfluidics will be emphasized. Finally, conditions for the spreading of microanalytical systems in routine proteomic labs will be discussed.

Electrophoresis↗

PubMed2003

Protein fractionation in a multicompartment device using Off-Gel isoelectric focusing.

A new protein fractionation technique based on off-gel isoelectric focusing (IEF) is presented, where the proteins are separated according to their isoelectric point (pI) in a multiwell device with the advantage to be directly recovered in solution for further analysis. The protein fractions obtained with this technique have then been characterized with polymer nanoelectrospray for mass spectrometry (MS) analyses or with Bioanalyzer for mass identification. This methodology shows the possibility of developing alternatives to the classical two-dimensional (2-D) gel electrophoresis. One species numerical simulation of the electric field distribution during off-gel separation is also presented in order to demonstrate the principle of the purification. Experiments with pI protein markers have been carried out in order to highlight the kinetics and the efficiency of the technique. Moreover, the resolution of the fractionation was shown to be 0.1 pH unit for the separation of beta-lactoglobulin A and B. In addition, the isoelectric fractionation of an Escherichia coli extract was performed in standard solubilization buffer to demonstrate the performances of the technique, notably for proteomics applications.

Animals↗

PubMed2002

Finite element simulation of pinched pressure-driven flow injection in microchannels.

A pinched pressure-driven flow injection on a microchip is numerically simulated in order to optimize the relative values of the operational parameters. The geometry studied is a two-dimensional rectangular channel featuring a cross-junction with a large depth-over-width ratio. The hydrodynamic and convection-diffusion equations are solved for the two steps of the process: first, the sample solution is pinched into the transversal channel (injection channel), and then it is injected into the longitudinal one (separation channel), where the time evolution of the concentration is analyzed for different types of the detectors. Electroosmotic flow calculations have also been performed and have shown a good agreement with literature. The results for pressure-driven flow point out that the shape of the detection signal is strongly dependent on the velocity in the separation channel and on the position of the detection probes. The so-called double-humped peak, caused by the parabolic flow profile at high driving flow rate is analyzed. A tight pinch greatly decreases the amount of injected sample and, consequently, the signal sensitivity without increasing its quality. A proper pullback of the sample during the separation process can decrease the tailing due to the sample leakage from the injection channel. Although a high sample pullback causes a considerable decrease in the signal sensitivity, it also greatly enhances the peak resolution. Finally, it is shown that a wider injection channel with high sample pullback ensures an improved signal sensitivity with good resolution.

Journal Article↗

PubMed2002

Mixing processes in a zigzag microchannel: finite element simulations and optical study.

A finite element model has been used in order to study the mixing process of species in a 100-microm-wide zigzag microchannel integrating a "Y" inlet junction. The distribution of the concentration was obtained by solving successively the Navier-Stokes equation and the diffusion-convection equation in the steady state form. Because of the large range of Reynolds numbers studied (1 < Re < 800), the 2D diffusion-convection simulations are carried out with high diffusion coefficients. The results illustrated the effects of both flow rate and channel geometry on hydrodynamics and mixing efficiency. Below a critical Reynolds number of approximately 80, the mixing is entirely ensured by molecular diffusion. For higher Reynolds numbers, simulations revealed the mixing contribution of laminar flow recirculations. This effect increases for lower values of diffusion coefficients. Experimental studies on the mixing of species at different flow rates are reported showing the same hydrodynamic tendency.

Equipment Design↗

PubMed2002

Finite element simulation of Off-Gel trade mark buffering.

The protonation of an aqueous solution of two ampholytes AH and BH next to a gel buffered by immobilized acid moieties IH has been studied by finite element simulation in an iterative scheme. A ten species model has been formulated, taking into account transient diffusion and equilibrium kinetics of the two amphoteric species AH and BH, of water and of the immobilized species IH. This model has been developed to illustrate the pH evolution between an ampholyte solution and an Immobiline gel, and to study the influence of the Immobiline concentration on protons and ampholyte distributions. It has been demonstrated that a minimum initial Immobiline concentration of 10(-2) M is necessary to maintain the pH in the gel in contact with a closed chamber, when the difference between the isoelectric points of AH and BH is 4 and when the initial concentration of the ampholytes in solution is in the micromolar range. This approach provides a first theoretical framework for the recently developed Off-Gel trade mark electrophoresis.

Buffers↗