Université PSL

Publications

RECHERCHER

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A new microfluidic approach for the one-step capture, amplification and label-free quantification of bacteria from raw samples
Laboratoire Macromolécules et Microsystèmes en Biologie et Médecine - Iago Pereiro, Amel Bendali, Sanae Tabnaoui, Lucile Alexandre, Jana Srbova, Zuzana Bilkova, Shane Deegan, Lokesh Joshi, Jean-Louis Viovy, Laurent Malaquin, Bruno Dupuy and Stéphanie Descroix
Chem. Sci. - 8(2) 1329-1336 - DOI: 10.1039/C6SC03880H - 2019
A microfluidic method to specifically capture and detect infectious bacteria based on immunorecognition and proliferative power is presented. It involves a microscale fluidized bed in which magnetic and drag forces are balanced to retain antibody-functionalized superparamagnetic beads in a chamber during sample perfusion. Captured cells are then cultivated in situ by infusing nutritionally-rich medium. The system was validated by the direct one-step detection of Salmonella Typhimurium in undiluted unskimmed milk, without pre-treatment. The growth of bacteria induces an expansion of the fluidized bed, mainly due to the volume occupied by the newly formed bacteria. This expansion can be observed with the naked eye, providing simple low-cost detection of only a few bacteria and in a few hours. The time to expansion can also be measured with a low-cost camera, allowing quantitative detection down to 4 cfu (colony forming unit), with a dynamic range of 100 to 107 cfu ml−1 in 2 to 8 hours, depending on the initial concentration. This mode of operation is an equivalent of quantitative PCR, with which it shares a high dynamic range and outstanding sensitivity and specificity, operating at the live cell rather than DNA level. Specificity was demonstrated by controls performed in the presence of a 500× excess of non-pathogenic Lactococcus lactis. The system's versatility was demonstrated by its successful application to the detection and quantitation of Escherichia coli O157:H15 and Enterobacter cloacae. This new technology allows fast, low-cost, portable and automated bacteria detection for various applications in food, environment, security and clinics.
Magnetic fluidized bed for solid phase extraction in microfluidic systems
Laboratoire Macromolécules et Microsystèmes en Biologie et Médecine - Pereiro, Iago ; Tabnaoui, Sanae ; Fermigier, Marc ; du Roure, Olivia ; Descroix, Stephanie ; Viovy, Jean-Louis ; Malaquin, Laurent
Lab. Chip - 17, 9 1603-1615 - DOI: 10.1039/C7LC00063D - 2019
Fluidization, a process in which a granular solid phase behaves like a fluid under the influence of an imposed upward fluid flow, is routinely used in many chemical and biological engineering applications. It brings, to applications involving fluid–solid exchanges, advantages such as high surface to volume ratio, constant mixing, low flow resistance, continuous operation and high heat transfer. We present here the physics of a new miniaturized, microfluidic fluidized bed, in which gravity is replaced by a magnetic field created by an external permanent magnet, and the solid phase is composed of magnetic microbeads with diameters ranging from 1 to 5 μm. These beads can be functionalized with different ligands, catalysts or enzymes, in order to use the fluidized bed as a continuous purification column or bioreactor. It allows flow-through operations at flow rates ranging from 100 nL min−1 up to 5 μL min−1 at low driving pressures (<100 mbar) with intimate liquid/solid contact and a continuous recirculation of beads for enhanced target capture efficiencies. The physics of the system presents significant differences as compared to conventional fluidized beds, which are studied here. The effects of magnetic field profile, flow chamber shape and magnetic bead dipolar interactions on flow regimes are investigated, and the different regimes of operation are described. Qualitative rules to obtain optimal operation are deduced. Finally, an exemplary use as a platform for immunocapture is provided, presenting a limit of detection of 0.2 ng mL−1 for 200 μL volume samples.
The power of solid supports in multiphase and droplet-based microfluidics: towards clinical applications
Laboratoire Macromolécules et Microsystèmes en Biologie et Médecine - Serra, M; Ferraro, D; Pereiro, I; Viovy, J-L; Descroix, S
Lab. Chip - 17 3979-3999 - DOI:10.1039/c7lc00582b - 2019
Multiphase and droplet microfluidic systems are growing in relevance in bioanalytical-related fields, especially due to the increased sensitivity, faster reaction times and lower sample/reagent consumption of many of its derived bioassays. Often applied to homogeneous (liquid/liquid) reactions, innovative strategies for the implementation of heterogeneous (typically solid/liquid) processes have recently been proposed. These involve, for example, the extraction and purification of target analytes from complex matrices or the implementation of multi-step protocols requiring efficient washing steps. To achieve this, solid supports such as functionalized particles (micro or nanometric) presenting different physical properties (e.g. magnetic, optical or others) are used for the binding of specific entities. The manipulation of such supports with different microfluidic principles has both led to the miniaturization of existing biomedical protocols and the development of completely new strategies for diagnostics and research. In this review, multiphase and droplet-based microfluidic systems using solid suspensions are presented and discussed with a particular focus on: i) working principles and technological developments of the manipulation strategies and ii) applications, critically discussing the level of maturity of these systems, which can range from initial proofs of concept to real clinical validations.
Chromatin immunoprecipitation in microfluidic droplets: towards fast and cheap analyses
Laboratoire Macromolécules et Microsystèmes en Biologie et Médecine - Teste, Bruno, Champ, Jerome ; Londono-Vallejo, Arturo, Descroix, Stephanie, Malaquin, Laurent, Viovy, Jean-Louis, Draskovic, Irena, Mottet, Guillaume
Lab. Chip - 17, 3: 530-537 - DOI: 10.1039/c6lc01535b - 2017
Genetic organization is governed by the interaction of DNA with histone proteins, and differential modifications of these proteins is a fundamental mechanism of gene regulation. Histone modifications are primarily studied through chromatin immunoprecipitation (ChIP) assays, however conventional ChIP procedures are time consuming, laborious and require a large number of cells. Here we report for the first time the development of ChIP in droplets based on a microfluidic platform combining nanoliter droplets, magnetic beads (MB) and magnetic tweezers (MT). The droplet approach enabled compartmentalization and improved mixing, while reducing the consumption of samples and reagents in an integrated workflow. Anti-histone antibodies grafted to MB were used as a solid support to capture and transfer the target chromatin from droplets to droplets in order to perform chromatin immunoprecipitation, washing, elution and purification of DNA. We designed a new ChIP protocol to investigate four different types of modified histones with known roles in gene activation or repression. We evaluated the performances of this new ChIP in droplet assay in comparison with conventional methods. The proposed technology dramatically reduces analytical time from a few days to 7 hours, simplifies the ChIP protocol and decreases the number of cells required by 100 fold while maintaining a high degree of sensitivity and specificity. Therefore this droplet-based ChIP assay represents a new, highly advantageous and convenient approach to epigenetic analyses.
FISH-in-CHIPS: A Microfluidic Platform for Molecular Typing of Cancer Cells.
Laboratoire Macromolécules et Microsystèmes en Biologie et Médecine - Serra, M ; Pereiro, I; Yamada, A; Viovy, J. –L.; Descroix, S. Ferraro
Methods Mol Biol. - 1547 211-220 - doi: 10.1007/978-1-4939-6734-6_16 - 2017
The sealing of microfluidic devices remains a complex and time-consuming process requiring specific equipment and protocols: a universal method is thus highly desirable. We propose here the use of a commercially available sealing tape as a robust, versatile, reversible solution, compatible with cell and molecular biology protocols, and requiring only the application of manually achievable pressures. The performance of the seal was tested with regards to the most commonly used chip materials. For most materials, the bonding resisted 5 bars at room temperature and 1 bar at 95 °C. This method should find numerous uses, ranging from fast prototyping in the laboratory to implementation in low technology environments or industrial production.
A simple and low-cost chip bonding solution for high pressure, high temperature and biological applications Serra, M ; Pereiro, I; Yamada, A; Viovy, J. –L.; Descroix, S. Ferraro
Laboratoire Macromolécules et Microsystèmes en Biologie et Médecine - Serra, M ; Pereiro, I; Yamada, A; Viovy, J. –L.; Descroix, S. Ferraro
Lab. Chip - 0.71111111111 629-634 - doi: 10.1039/c6lc01319h - 2017
The sealing of microfluidic devices remains a complex and time-consuming process requiring specific equipment and protocols: a universal method is thus highly desirable. We propose here the use of a commercially available sealing tape as a robust, versatile, reversible solution, compatible with cell and molecular biology protocols, and requiring only the application of manually achievable pressures. The performance of the seal was tested with regards to the most commonly used chip materials. For most materials, the bonding resisted 5 bars at room temperature and 1 bar at 95 °C. This method should find numerous uses, ranging from fast prototyping in the laboratory to implementation in low technology environments or industrial production.
Droplet Microfluidic and Magnetic Particles Platform for Cancer Typing.
Laboratoire Macromolécules et Microsystèmes en Biologie et Médecine - Ferraro D, Champ J, Teste B, Serra M, Malaquin L, Descroix S, de Cremoux P, Viovy JL.
Lab. Chip - 1547 113-121 - doi: 10.1007/978-1-4939-6734-6_9. - 2017
Analyses of nucleic acids are routinely performed in hospital laboratories to detect gene alterations for cancer diagnosis and treatment decision. Among the different possible investigations, mRNA analysis provides information on abnormal levels of genes expression. Standard laboratory methods are still not adapted to the isolation and quantitation of low mRNA amounts and new techniques needs to be developed in particular for rare subsets analysis. By reducing the volume involved, time process, and the contamination risks, droplet microfluidics provide numerous advantages to perform analysis down to the single cell level.We report on a droplet microfluidic platform based on the manipulation of magnetic particles that allows the clinical analysis of tumor tissues. In particular, it allows the extraction of mRNA from the total-RNA sample, Reverse Transcription, and cDNA amplification, all in droplets.
Transient microfluidic compartmentalization using actionable microfilaments for biochemical assays, cell culture and organs-on-chip.
Laboratoire Macromolécules et Microsystèmes en Biologie et Médecine - Yamada A, Renault R, Chikina A, Venzac B, Pereiro I, Coscoy S, Verhulsel M, Parrini MC, Villard C, Viovy JL, Descroix S.
Lab. Chip - 16(24) 16(24) - DOI: 10.1039/c6lc01143h - 2016
We report here a simple yet robust transient compartmentalization system for microfluidic platforms. Cylindrical microfilaments made of commercially available fishing lines are embedded in a microfluidic chamber and employed as removable walls, dividing the chamber into several compartments. These partitions allow tight sealing for hours, and can be removed at any time by longitudinal sliding with minimal hydrodynamic perturbation. This allows the easy implementation of various functions, previously impossible or requiring more complex instrumentation. In this study, we demonstrate the applications of our strategy, firstly to trigger chemical diffusion, then to make surface co-coating or cell co-culture on a two-dimensional substrate, and finally to form multiple cell-laden hydrogel compartments for three-dimensional cell co-culture in a microfluidic device. This technology provides easy and low-cost solutions, without the use of pneumatic valves or external equipment, for constructing well-controlled microenvironments for biochemical and cellular assays.
Asymmetric axonal edge guidance: a new paradigm for building oriented neuronal networks.
Laboratoire Macromolécules et Microsystèmes en Biologie et Médecine - Renault R, Durand JB, Viovy JL, Villard C.
Lab. Chip - 16(12) 2188-91 - doi: 10.1039/c6lc00479b - 2016
We present a novel kind of directional axon guides for brain-on-a-chip applications. Contrarily to previous works, the directionality in our design is created by rerouting axons growing in the unwanted direction back to their original compartment while leaving the other growth direction unaffected. This design yields state-of-the-art levels of directionality without the disadvantages of previously reported technologies.
In-mold patterning and actionable axo-somatic compartmentalization for on-chip neuron culture.
Laboratoire Macromolécules et Microsystèmes en Biologie et Médecine - Yamada A, Vignes M, Bureau C1, Mamane A, Venzac B, Descroix S, Viovy JL, Villard C, Peyrin JM, Malaquin L.
Lab. Chip - 16(11) 2059-68 - doi: 10.1039/c6lc00414h. - 2016
Oriented neuronal networks with controlled connectivity are required for many applications ranging from studies of neurodegeneration to neuronal computation. To build such networks in vitro, an efficient, directed and long lasting guidance of axons toward their target is a pre-requisite. The best guidance achieved so far, however, relies on confining axons in enclosed microchannels, making them poorly accessible for further investigation. Here we describe a method providing accessible and highly regular arrays of axons, emanating from somas positioned in distinct compartments. This method combines the use of a novel removable partition, allowing soma positioning outside of the axon guidance patterns, and in-mold patterning (iMP), a hybrid method combining chemical and mechanical cell positioning clues applied here for the first time to neurons. The axon guidance efficiency of iMP is compared to that of conventional patterning methods, e.g. micro-contact printing (chemical constraints by a poly-l-lysine motif) and micro-grooves (physical constraints by homogeneously coated microstructures), using guiding tracks of different widths and spacing. We show that iMP provides a gain of 10 to 100 in axon confinement efficiency on the tracks, yielding mm-long, highly regular, and fully accessible on-chip axon arrays. iMP also allows well-defined axon guidance from small populations of several neurons confined at predefined positions in μm-sized wells. iMP will thus open new routes for the construction of complex and accurately controlled neuronal networks.

28 publications.