Impedimetric array in polymer microfluidic cartridge for low cost point-of-care diagnostics

International audienc

Elaboration de nouveaux biocapteurs électrochimiques pour le diagnostic de la thrombose veineuse profonde

La thrombose veineuse profonde (TVP) comme sous le nom de phlébite correspond à la formation d'un caillot sanguin dans une veine. C'est une pathologie fréquente en Europe et elle reste très difficile à diagnostiquer jusqu'à présent. Ce travail qui rentre dans le cadre d'un projet Européen STREP C] consiste à élaborer un biomatériau en tant qu'outil de diagnostic de la thrombose permettant une mesure en temps réel de la présence de la maladie dans le sang du patient. La détection est réalisée par dosage de la glycoprotéine D-Dimère présente lors de la formation d'un thrombus. L'approche qui a été envisagée dans cette étude est celle de l'immobilisation de l'anticorps anti-D-Dimère tagué Histidine sur un matériau polymérique conducteur fonctionnalisé par un complexe métallique. Cette stratégie originale de la construction de l'immunocapteur réalisée étape par étape a permis à la fois d'avoir une fixation orientée de l'anticorps et de suivre la variation du signal électrochimique via le métal utilisé en tant que sonde redox. De ce fait un biocapteur a été développé à base de polypyrrole fonctionnalisé par le complexe métallique Acide NitrilotriacétiquelMétal ionique (NT AJCu2+) et a été intégré par la suite dans un dispositif miniaturisé conçu pour être fiable, précis, portatif, automatisé et économique. L'étude de stabilité dans le temps de ce biomatériau a permis la réalisation d'une nouvelle matrice d'immobilisation de l'anticorps anti-D-Dimère en utilisant un nouveau ligand enPI2 constitué par des groupements amines comms pour une très bonne chélation du cuivre II. La conception du biocapteur sur des nanomatériaux tels que les nanotubes de carbone et les nanoparticules d'or a montré une augmentation de la gamme de détection du D-Dimère. La caractérisation de ces biomatériaux et le suivi du D-Dimère ont nécessité l'utilisation de différentes techniques d'analyses électrochimique et de surface tels que la voltammétrie cyclique (CV), la DPV, la spectroscopie d'impédance électrochimique (SIE), la microscopie à force atomique (AFM), la résonance des plasmons de surface (SPR), L'infrarouge à transformée de Fourrier (FTIR), la spectroscopie des photoélectrons X (XPS) et la résonance magnétique nucléaire (RPE). L'intégration du biocapteur dans la cartouche DVT-IMP comprenant une puce et un corps micro fluidique a permis l'amplification du signal électrochimique notamment de la sonde redox grâce à l'utilisation des élecctrodes interdigitées (EIDs).Deep vein thrombosis (DVT) known as phlebitis corresponds to the formation of a blood clot in a vein. It is a common disorder in Europe and it remains very difficult to diagnose until now. This work within the framework of a European project STREP [] is to develop a biomaterial as a tool for diagnosis of thrombosis allowing real-time measurement of the presence of the disease in the patient's blood. Detection is perfonned by assaying the glycoprotein D-dimer present during the formation of a thrombus. The approach which has been chosen in this study is the immobilization of an Histidine tagged anti-D-Dimer on a conductive polymer material functionalized with a metal complex. This original strategy for the construction of the immunosensor has been achieved step by step allowed both to have an oriented attachment of the antibody and monitor the change in electrochemical signal through the metal used as a redox probe. Thus a biosensor based on polypyrrole functionalized with nitrilotriacetic acid metal complex (NT AJCu2 +) was developed and was subsequently incorporated into a miniaturized device designed to be reliable, accurate, portable, automated and economical. The study of the biomaterial time stability has enabled the creation of a new matrix for the anti-D-dimer immobilization using a new ligand enPI2 containing amine groups known for a very good Copper II chelation. The design of the biosensor on nanomaterials such as carbon nanotubes and gold nanoparticles showed an increase in the D-dimer detection range. The characterization of these biomaterials and the D-dimer monitoring required the use of different electrochemical and surface analytical techniques such as cyclic voltammetry (CV), DifferentiaI Pulse Voltammetry (DPV), the electrochemical impedance spectroscopy (EIS), the Atomic force microscopy (AFM), the surface plasmon resonance (SPR), The Fourier transform infrared (FTIR), X-ray photoelectron spectroscopy (XPS) and nuclear paramagnetic resonance (EPR). The integration of the biosensor in the DVT-IMP cartridge comprising a chip and a microfluidic body allowed the amplification of the electrochemical signal including the redox probe through the use of interdigitated élecctrodes (EIDs).ORSAY-PARIS 11-BU Sciences (914712101) / SudocSudocFranceF

Determination of the isomeric forms proportion of fluorogenic naphthalene-2,3-dicarboxaldehyde in a binary mixture of water:methanol using electrochemical methods.

International audienceThe electrochemical response of the fluorogenic label naphthalene-2,3-dicarboxyaldehyde (NDA) in a binary mixture of water/methanol was characterized with cyclic voltammetry (CV) and differential pulse voltammetry (DPV) electrochemical techniques. Naphthalene-2,3-dicarboxyaldehyde does exist in three isomeric forms in aqueous solution: the unhydrated dialdehyde (DA), the acyclic monohydrated (MA) and the cyclic hemiacetal (HAC). The study underlines that the proportion of each of them varies according to the working pH. At low and high pH, the dialdehyde form is in larger proportion than the acyclic monohydrated form. Conversely at intermediate pH, the concentration of the acyclic form is in greater proportion than the dialdehyde form. These results allowed us to determine the optimal pH of 9 for which the labeling of biomolecules could be more efficient due to the base catalyzed regeneration of the unhydrated form. At this pH, the data processing from the analysis of measured currents and estimation of diffusion coefficients of each form according to the semi-empirical models of Wilke-Chang, Scheibel, Reddy-Doraiswamy and Lusis-Ratcliff allowed us to obtain the concentration of dialdehyde (0.28mM), acyclic monohydrated (0.57mM) and cyclic hemiacetal monohydrated (0.15mM) forms starting from 1mM naphthalene-2,3-dicarboxyaldehyde

Improved electrochemical detection of a transthyretin synthetic peptide in the nanomolar range with a two-electrode system integrated in a glass/PDMS microchip.

International audienceAn alternative to a three-electrode set-up for electrochemical detection and analysis in microfluidic chips is described here. The design of the electrochemical sensor consists of the surface of the glass substrate covered with a PDMS block which bears the microfluidic channels. A band microelectrode which acts as a working electrode surrounded by a large counter electrode is obtained at the micrometric level to propose a simple and efficient sensing area for on-a-chip analysis. The counter-electrode with a surface area about 22-fold greater than the working-microelectrode can also be considered as a pseudo reference since its current density is low and thus limits the potential variations around the rest potential. To this purpose, the [Fe(iii)(CN)6](3-)/[Fe(ii)(CN)6](4-) redox couple was used in order to set a reference potential at 0 V since both electrodes were platinum. The electrochemical microchip performance was characterized using differential pulse voltammetric (DPV) detection and quantification of the optically multi-labelled transthyretin synthetic peptide mimicking a tryptic fragment of interest for the diagnosis of familial transthyretin amyloidosis (ATTR). The limit of detection of the peptide by the working microelectrode was 25 nM, a value 100-fold lower than the one reported with conventional capillary electrophoresis coupled with laser-induced fluorescence under the same analytical conditions

Study of Surface Charge Instabilities by EOF Measurements on a Chip: A Real-Time Hysteresis and Peptide Adsorption Based Methodology.

International audienceThis paper describes the measurement of the electroosmotic mobility (EOF) in a Wheatstone fluidic bridge (μFWB) as a direct probe of the surface instability. The variation of EOF known as one major contribution of the electrokinetic migration has been determined with a real-time measurement platform after different conditionings on chips. We also scan the pH of the background electrolytes with three different ionic strengths to evaluate the dependencies of the EOF as a function of the pH. A hysteresis methodology has been developed for probing the surface charge instabilities. EOF mobility has been recorded during on-a-chip electrophoresis to estimate the effect of such instability on the analytical performance. As expected, our experimental curves show that a decrease in the ionic strength increases the surface charge stability of the hybrid microchip. This result demonstrates that ionic exchanges between the surface and the fluid are clearly involved in the stability of the surface charge. With this original method based on real-time EOF measurement, the surface state can be characterized after hydrodynamic and electrophoresis sequences to mimic any liquid conditioning and separation steps. Finally, as a demonstrative application, isotherms of the adsorption of insulin have been recorded showing the change in surface charge by unspecific adsorption of this biomolecule onto the microfluidic channel's wall. These methodologies and findings could be particularly relevant to investigating various analytical pathways and to understanding the molecular mechanisms at solid/liquid interfaces