Université PSL

Publications

RECHERCHER

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Single-cell deep phenotyping of IgG-secreting cells for high-resolution immune monitoring.
Eyer K, Doineau RCL,Castrillon C, Briseño-Roa L, Menrath V, Mottet G, England P, Godina A, Brient-Litzler E, Nizak C, Jensen A, Griffiths AD, Bibette J, Bruhns P, Baudry J.
Nat Biotechnol. - 35(10) 977-982 - doi: 10.1038/nbt.3964. -
Studies of the dynamics of the antibody-mediated immune response have been hampered by the absence of quantitative, high-throughput systems to analyze individual antibody-secreting cells. Here we describe a simple microfluidic system, DropMap, in which single cells are compartmentalized in tens of thousands of 40-pL droplets and analyzed in two-dimensional droplet arrays using a fluorescence relocation-based immunoassay. Using DropMap, we characterized antibody-secreting cells in mice immunized with tetanus toxoid (TT) over a 7-week protocol, simultaneously analyzing the secretion rate and affinity of IgG from over 0.5 million individual cells enriched from spleen and bone marrow. Immunization resulted in dramatic increases in the range of both single-cell secretion rates and affinities, which spanned at maximum 3 and 4 logs, respectively. We observed differences over time in dynamics of secretion rate and affinity within and between anatomical compartments. This system will not only enable immune monitoring and optimization of immunization and vaccination protocols but also potentiate antibody screening.
Hydrophobization of Silica Nanoparticles in Water: Nanostructure and Response to Drying Stress
Solenn Moro, Caroline Parneix, Bernard Cabane, Nicolas Sanson , and Jean-Baptiste d’Espinose de Lacaillerie
Langmuir - 33 (19) 4709–4719 - DOI: 10.1021/acs.langmuir.6b04505 -
We report on the impact of surface hydrophobization on the structure of aqueous silica dispersions and how this structure resists drying stress. Hydrophilic silica particles were hydrophobized directly in water using a range of organosilane precursors, with a precise control of the grafting density. The resulting nanostructure was precisely analyzed by a combination of small-angle X-ray scattering (SAXS) and cryo-microscopy (cryo-TEM). Then, the dispersion was progressively concentrated by drying, and the evolution of the nanostructures as a function of the grafting density was followed by SAXS. At the fundamental level, because the hydrophobic character of the silica surfaces could be varied continuously through a precise control of the grafting density, we were able to observe how the hydrophobic interactions change particles interactions and aggregates structures. Practically, this opened a new route to tailor the final structure, the residual porosity, and the damp-proof properties of the fully dried silica. For example, regardless of the nature of the hydrophobic precursor, a grafting density of 1 grafter per nm2 optimized the interparticle interactions in solution in view to maximize the residual porosity in the dried material (0.9 cm3/g) and reduced the water uptake to less than 4% in weight compared to the typical value of 13% for hydrophilic particles (at T = 25 °C and relative humidity = 80%).
The mechanism of eccrine sweat pore plugging by aluminium salts using microfluidics combined with small angle X-ray scattering
Alice Bretagne, Franck Cotot, Mireille Arnaud-Roux, Michael Sztucki, Bernard Cabane and Jean-Baptiste Galey
Soft Matter - 13 3812-3821 - 10.1039/C6SM02510B -
Aluminium salts are widely used to control sweating for personal hygiene purposes. Their mechanism of action as antiperspirants was previously thought to be a superficial plugging of eccrine sweat pores by the aluminium hydroxide gel. Here we present a microfluidic T junction device that mimics sweat ducts, and is designed for the real time study of interactions between sweat and ACH (Aluminium Chloro Hydrate) under conditions that lead to plug formation. We used this device to image and measure the diffusion of aluminium polycationic species in sweat counter flow. We report the results of small angle X-ray scattering experiments performed to determine the structure and composition of the plug, using BSA (Bovine Serum Albumin) as a model of sweat proteins. Our results show that pore occlusion occurs as a result of the aggregation of sweat proteins by aluminium polycations. Mapping of the device shows that this aggregation is initiated in the T junction at the location where the flow of aluminium polycations joins the flow of BSA. The mechanism involves two stages: (1) a nucleation stage in which aggregates of protein and polycations bind to the wall of the sweat duct and form a tenuous membrane, which extends across the junction; (2) a growth stage in which this membrane collects proteins that are carried by hydrodynamic flow in the sweat channel and polycations that diffuse into this channel. These results could open up perspectives to find new antiperspirant agents with an improved efficacy.
Interparticle Capillary Forces at a Fluid–Fluid Interface with Strong Polymer-Induced Aging
Stefano Cappelli , Arthur M. de Jong, Jean Baudry, and Menno W. J. Prins
Langmuir - 33 (3) 696–705 - DOI: 10.1021/acs.langmuir.6b03910 -
We report on a measurement of forces between particles adsorbed at a water–oil interface in the presence of an oil-soluble polymer. The cationic polymer interacts electrostatically with the negatively charged particles, thereby modulating the particle contact angle and the magnitude of capillary attraction between the particles. However, polymer adsorption to the interface also generates an increase in the apparent interfacial viscosity over several orders of magnitude in a time span of a few hours. We have designed an experiment in which repeated motion trajectories are measured on pairs of particles. The experiment gives an independent quantification of the interfacial drag coefficient (10–7–10–4 Ns/m) and of the interparticle capillary forces (0.1–10 pN). We observed that the attractive capillary force depends on the amount of polymer in the oil phase and on the particle pair. However, the attraction appears to be independent of the surface rheology, with changes over a wide range of apparent viscosity values due to aging. Given the direction (attraction), the range (∼μm), and the distance dependence (∼1/S5) of the observed interparticle force, we interpret the force as being caused by quadrupolar deformations of the fluid–fluid interface induced by particle surface roughness. The results suggest that capillary forces are equilibrated in the early stages of interface aging and thereafter do not change anymore, even though strong changes in surface rheology still occur. The described experimental approach is powerful for studying dissipative as well as conservative forces of micro- and nanoparticles at fluid–fluid interfaces for systems out of equilibrium.
Controlled production of sub-millimeter liquid core hydrogel capsules for parallelized 3D cell culture
Hugo Doméjean, Mathieu de la Motte Saint Pierre, Anette Funfak, Nicolas Atrux-Tallau, Kevin Alessandri, Pierre Nassoy, Jérôme Bibette and Nicolas Bremond
Lab. Chip - 17 110-119 - DOI: 10.1039/C6LC00848H -
Liquid core capsules having a hydrogel membrane are becoming a versatile tool for three-dimensional culture of micro-organisms and mammalian cells. Making sub-millimeter capsules at a high rate, via the breakup of a compound jet in air, opens the way to high-throughput screening applications. However, control of the capsule size monodispersity, especially required for quantitative bioassays, was still lacking. Here, we report how the understanding of the underlying hydrodynamic instabilities that occur during the process can lead to calibrated core–shell bioreactors. The requirements are: i) damping the shear layer instability that develops inside the injector arising from the co-annular flow configuration of liquid phases having contrasting viscoelastic properties; ii) controlling the capillary instability of the compound jet by superposing a harmonic perturbation onto the shell flow; iii) avoiding coalescence of drops during jet fragmentation as well as during drop flight towards the gelling bath; iv) ensuring proper engulfment of the compound drops into the gelling bath for building a closed hydrogel shell. We end up with the creation of numerous identical compartments in which cells are able to form multicellular aggregates, namely spheroids. In addition, we implement an intermediate composite hydrogel layer, composed of alginate and collagen, allowing cell adhesion and thus the formation of epithelia or monolayers of cells.
Osmotic pressures of lysozyme solutions from gas-like to crystal states
Coralie Pasquier,ab Sylvie Beaufils,b Antoine Bouchoux, Sophie Rigault, Bernard Cabane, Mikael Lund, Valérie Lechevalier,a Cécile Le Floch-Fouéré,a Maryvonne Pasco,a Gilles Pabœuf,b Javier Pérezf and Stéphane Pezennec*a
Phys. Chem. - 18 28458-28465 - DOI: 10.1039/C6CP03867K -
We obtained osmotic pressure data of lysozyme solutions, describing their physical states over a wide concentration range, using osmotic stress for pressures between 0.05 bar and about 40 bar and volume fractions between 0.01 and 0.61. The osmotic pressure vs. volume fraction data consist of a dilute, gas-phase regime, a transition regime with a high-compressibility plateau, and a concentrated regime where the system is nearly incompressible. The first two regimes are shifted towards a higher protein volume fraction upon decreasing the strength or the range of electrostatic interactions. We describe this shift and the overall shape of the experimental data in these two regimes through a model accounting for a steric repulsion, a short-range van der Waals attraction and a screened electrostatic repulsion. The transition is caused by crystallization, as shown by small-angle X-ray scattering. We verified that our data points correspond to thermodynamic equilibria, and thus that they consist of the reference experimental counterpart of a thermodynamic equation of state.
Coarse-grained modeling of the intrinsically disordered protein Histatin 5 in solution: Monte Carlo simulations in combination with SAXS.
Cragnell C, Durand D, Cabane B, Skepö
Proteins - 84(6) 777-91 - doi: 10.1002/prot.25025. -
Monte Carlo simulations and coarse-grained modeling have been used to analyze Histatin 5, an unstructured short cationic salivary peptide known to have anticandidical properties. The calculated scattering functions have been compared with intensity curves and the distance distribution function P(r) obtained from small angle X-ray scattering (SAXS), at both high and low salt concentrations. The aim was to achieve a molecular understanding and a physico-chemical insight of the obtained SAXS results and to gain information of the conformational changes of Histatin 5 due to altering salt content, charge distribution, and net charge. From a modeling perspective, the accuracy of the electrostatic interactions are of special interest. The used coarse-grained model was based on the primitive model in which charged hard spheres differing in charge and in size represent the ionic particles, and the solvent only enters the model through its relative permittivity. The Hamiltonian of the model comprises three different contributions: (i) excluded volumes, (ii) electrostatic, and (iii) van der Waals interactions. Even though the model can be considered as gross omitting all atomistic details, a great correspondence is obtained with the experimental results. Proteins 2016; 84:777-791.
Traffic collision during the breakup of an aqueous viscous compound jet
Hugo Doméjean, Jérôme Bibette, and Nicolas Bremond
Phys. Rev. Fluids - 1 063903 - https://doi.org/10.1103/PhysRevFluids.1.063903 -
Liquid jets ultimately break up into droplets through an instability driven by surface tension. For highly viscous liquids, drops are connected by cylindrical liquid filaments whose radii linearly decrease with time, thus forming drops on a string structure. For a jet composed of two aqueous phases made in air by coaxial extrusion, we observe that, for moderate Weber and capillary numbers, drops slow down with different velocities, leading to drop coalescence. The origin of the traffic collision is linked to the spatial feature of the capillary instability where capillary and viscous forces acting on the drops evolve along the jet and ultimately amplify small velocity fluctuations. The emergence of such fluctuations is related to the unstable nature of the annular coflow of liquids having contrasting viscoelastic properties. From a practical point of view, flow and actuation conditions can be adjusted to inhibit drop collision and thus drop coalescence. These findings allow then the fabrication of monodisperse submillimeter core-shell objects based on the fragmentation of compound jets made of polymer solutions that find applications for three-dimensional cell culture.
Microfluidic fabrication of composite hydrogel microparticles in the size range of blood cells
A. Pittermannová, Z. Ruberová, A. Zadražil, N. Bremond, J. Bibetteb and F. Štěpánek
RSC Adv. - 6 103532-103540 - DOI: 10.1039/C6RA23003B -
The fabrication of alginate hydrogel microparticles with embedded liposomes and magnetic nanoparticles for radiofrequency controlled release of encapsulated chemical cargo was considered. An extractive gelation process was implemented in a microfluidic device, which enabled the production of uniform composite microparticles of dimensions comparable to those of blood cells (between 5 and 10 μm). The critical parameters that control the extractive gelation process were systematically explored and feasible values that provide microgel particles of a defined size and morphology were identified. First, the initial water-in-oil droplet is formed in a flow-focusing junction whose size is controlled by the flow-rate of the oil phase. Then, the train of droplets is sandwiched between two streams of oil containing calcium ions. In that way, a flux of water molecules from the droplets towards the continuous phase as well as a transport of calcium ions towards the disperse phase are initiated. The final microparticle properties were thus found to be sensitive to three elementary sub-processes: (i) the initial droplet size; (ii) the extraction of water into the oil phase, which was controlled by the volume of the oil phase and its initial moisture content; and (iii) the kinetics of ionic cross-linking of the alginate matrix, which was controlled by the varying calcium concentration. The size and morphology of the final composite microgels were fully characterized.
Interfacial rheometry of polymer at a water–oil interface by intra-pair magnetophoresis
Stefano Cappelli, Arthur M. de Jong, Jean Baudryc and Menno W. J. Prins
Soft Matter - 12 5551-5562 - DOI: 10.1039/C5SM02917A -


We describe an interfacial rheometry technique based on pairs of micrometer-sized magnetic particles at a fluid–fluid interface. The particles are repeatedly attracted and repelled by well-controlled magnetic dipole–dipole forces, so-called interfacial rheometry by intra-pair magnetophoresis (IPM). From the forces (∼pN), displacements (∼μm) and velocities (∼μm s−1) of the particles we are able to quantify the interfacial drag coefficient of particles within a few seconds and over very long timescales. The use of local dipole–dipole forces makes the system insensitive to fluid flow and suited for simultaneously recording many particles in parallel over a long period of time. We apply IPM to study the time-dependent adsorption of an oil-soluble amino-modified silicone polymer at a water–oil interface using carboxylated magnetic particles. At low polymer concentration the carboxylated particles remain on the water side of the water–oil interface, while at high polymer concentrations the particles transit into the oil phase. Both conditions show a drag coefficient that does not depend on time. However, at intermediate polymer concentrations data show an increase of the interfacial drag coefficient as a function of time, with an increase over more than three orders of magnitude (10−7 to 10−4 N s m−1), pointing to a strong polymer-polymer interaction at the interface. The time-dependence of the interfacial drag appears to be highly sensitive to the polymer concentration and to the ionic strength of the aqueous phase. We foresee that IPM will be a very convenient technique to study fluid–fluid interfaces for a broad range of materials systems.

291 publications.