Naoufel Haddour

Microfluidic immunomagnetic cell separation using integrated permanent micromagnets

In this paper, we demonstrate the possibility to trap and sort labeled cells under flow conditions using a microfluidic device with an integrated flat micro-patterned hard magnetic film. The proposed technique is illustrated using a cell suspension containing a mixture of Jurkat cells and HEK (Human Embryonic Kidney) 293 cells. Prior to sorting experiments, the Jurkat cells were specifically labeled with immunomagnetic nanoparticles, while the HEK 293 cells were unlabeled. Droplet-based experiments demonstrated that the Jurkat cells were attracted to regions of maximum stray field flux density while the HEK 293 cells settled in random positions. When the mixture was passed through a polydimethylsiloxane (PDMS) microfluidic channel containing integrated micromagnets, the labeled Jurkat cells were selectively trapped under fluid flow, while the HEK cells were eluted towards the device outlet. Increasing the flow rate produced a second eluate much enriched in Jurkat cells, as revealed by flow cytometry. The separation efficiency of this biocompatible, compact micro-fluidic separation chamber was compared with that obtained using two commercial magnetic cell separation kits.

Characterization of C-PDMS electrodes for electrokinetic applications in microfluidic systems

This paper reports on the integration of thick carbon-polydimethylsiloxane (C-PDMS) electrodes in microfluidic systems for electrokinetic operations. The C-PDMS material, obtained by mixing carbon nanopowder and PDMS, preserves PDMS processing properties such as O2 plasma activation and soft-lithography patternability in thick or 3D electrodes. Conductivity in the order of 10 S m−1 was reached for a carbon concentration of 25 wt%. To evaluate the adhesion between PDMS and C-PDMS, we prepared bi-material strips and carried out a manual pull test. The cohesion and robustness of C-PDMS were also evaluated by applying a large range of electric field conditions from dc to ac (300 kHz). No damage to the electrodes or release of carbon was noticed. The use of such a material for electrokinetic manipulation was validated on polystyrene particles and cells. Here, we demonstrate that C-PDMS seems to be a valuable technological solution for electrokinetic in microfluidic and particularly for biological applications such as cell electrofusion, lysis and trapping, which are favored by uniform lateral electric fields across the microchannel section.

Micromagnet structures for magnetic positioning and alignment

High performance hard magnetic films (NdFeB, SmCo) have been patterned at the micron scale using thermo-magnetic patterning. Both out-of-plane and in-plane magnetized structures have been prepared. These micromagnet arrays have been used for the precise positioning and alignment of superparamagnetic nano- and microparticles. The specific spatial arrangement achieved is shown to depend on both the particle size and the size and orientation of the micromagnets. These micromagnet arrays were used to trap cells magnetically functionalized by endocytosis of 100 nm superparamagnetic particles. These simple, compact, and autonomous structures, which need neither an external magnetic field source nor a power supply, have much potential for use in a wide range of biological applications.

Microbial fuel cell anodic microbial population dynamics during MFC start-up

In order to optimize energy production in MFCs, a better understanding of anodic communities is essential. Our objective was to determine the taxonomic structure of the bacterial communities present at the surface of the anode during the formation and development of electro-active biofilms in MFCs inoculated with fresh primary clarifier overflow. Quantitative microbial community dynamics were evaluated as a function of time and electrical performance using 16S rRNA gene-based phylogenetic microarrays and flow cytometry. Results show that the bacterial community stabilized partially but not completely when voltage output was stable. Geobacter appeared to be the predominant genus, whose growth was associated with voltage, while some other genus still developed or declined after the voltage stabilization. Flow cytometry revealed that some genus showing a decreasing proportional fluorescence intensity over time were still actively respiring bacteria, and thus, active albeit minor members of the biofilm. Finally, this study shows that anodic biofilm selection and maturation is still occurring after more than 20 days of operation and over ten days after voltage is stabilized.

Trapping of Magnetically‐Labelled Liposomes on Flat Micro‐Patterned Hard Magnetic Films

In this paper, we describe the use of Giant Unilamellar Vesicles (GUV) as cellular models to illustrate the possibility to trap magnetically‐marked cells on a flat micro‐patterned hard magnetic film by magnetophoresis. GUV can encapsulate nano‐ or micro‐functional objects during the electroformation process and the amount of magnetic material incorporated can be controlled by regulating the concentration of magnetic nanoparticles in the electroformation buffer. The micromagnet arrays described in this paper are capable of generating high magnetic field gradients, which favors attraction of the magnetic GUV. These flat magnetic microstructures are obtained using the recently developed thermo‐magnetic patterning technique (TMP).

From Bipolar to Quadrupolar Electrode Structures: An Application of Bond-Detach Lithography for Dielectrophoretic Particle Assembly

We describe a new, simple process for fabricating transparent quadrupolar electrode arrays enabling large-scale particle assembly by means of dielectrophoresis. In the first step, interdigitated electrode arrays are made by chemical wet etching of indium tin oxide (ITO). Then, the transition from a bipolar to a quadrupolar electrode arrangement is obtained by covering the electrode surface with a thin poly(dimethylsiloxane) (PDMS) film acting as an electrical insulation layer in which selective openings are formed using bond-detach lithography. The PDMS insulating layer thickness was optimized and controlled by adjusting experimental parameters such as the PDMS viscosity (modulated by the addition of heptane) and the PDMS spin-coating velocity. The insulating character of the PDMS membrane was successfully demonstrated by performing a dielectrophoretic assembly of polystyrene particles using interdigitated electrodes with and without a PDMS layer. The results show that the patterned PDMS film functions properly as an electrical insulation layer and allows the reconfiguration of the electric field cartography. Electric field simulations were performed in both configurations to predict the dielectrophoretic behavior of the particles. The simulation results are in perfect agreement with experiments, in which we demonstrated the formation of concentrated clusters of polystyrene particles and living cells of regular size and shape.

Magnetic nanoparticle DNA labeling for individual bacterial cell detection and recovery

A culture independent approach was developed for recovering individual bacterial cells out of communities from complex environments including soils and sediments where autofluorescent contaminants hinder the use of fluorescence based techniques. For that purpose fifty nanometer sized streptavidin-coated superparamagnetic nanoparticles were used to chemically bond biotin-functionalized plasmid DNA molecules. We show that micromagnets can efficiently trap magnetically labeled transformed Escherichia coli cells after these bacteria were subjected to electro-transformation by these nanoparticle-labeled plasmids. Among other applications, this method could extend the range of approaches developed to study DNA dissemination among environmental bacteria without requiring cultivability of recombinant strains or expression of heterologous genes in the new hosts.

Tunable and label-free bacteria alignment using standing surface acoustic waves

This paper describes a new technique for focusing bacteria in a microfluidic channel and subsequently controlling their trajectory. Bacteria alignment is obtained using standing surface acoustic waves (SSAW) generated by two interdigitated transducer electrodes (IDTs) patterned on a piezoelectric wafer. The bacteria are focused in the standing wave pressure nodes, separated by half a wavelength, the electrode geometry and applied voltage frequency being chosen accordingly. Interestingly, the position of a pressure node can be modulated by introducing a phase shift between the electrical signals applied to both IDTs. The bacteria, trapped in this node, follow it and can therefore be deflected. This technique works with label-free bacteria in their culture medium and induces low power consumption, which is very interesting for portable devices.

Realization and characterization of carbonic layers on 4H-SiC for electrochemical detections

Carbon materials are considered building blocks for most of electrochemical sensors. Their biocompatibility allows their use as transducers for biosensors. Furthermore, they can be patterned, giving interest in all-inclusive bioelectrochemical detection devices. In particular, graphene, boron doped diamond, and pyrolyzed photoresist films are suitable for this kind of application, which would give the ability to use these working electrodes within a fully designed apparatus completed by counter and reference electrode. In this paper, a pioneer work is exposed on the synthesis of these materials for use as electrochemical sensors and as transducers for biodetection of acetylcholine by adsorption of acetylthiocholinesterase. The lowest limit of detection reaches 6.98 μM, sensitivity 5.91 μA/mM, and a linear range from 6.98 μM to 0.55 mM.

Electromagnetic characterization of biological cells

This paper presents the most commonly used method to characterize individual biological cells on a dielectric point of view. It is a force based technique which lays on dielectrophoresis and/or electrorotation. First the principle of these phenomena are described and analyzed with an extension to magnetic forces at the micrometric scale level. Secondly we present an experimental setup which permits to acquire the dielectrophoretic spectrum which is a dielectric signature of a cell. The main dielectric parameters can be deduced by fitting the theoretical response of the cell issued from a dielectric model and the experimental data. At the end we present an improved fitting method which takes advantage of a sensitivity analysis based on a probabilistic approach.