Université PSL



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Controlled proteolysis of normal and pathological prion protein in a microfluidic chip
Laboratoire Macromolécules et Microsystèmes en Biologie et Médecine - Le Nel A, Minc N, Smadja C, Slovakova M, Bilkova Z, Peyrin JM, Viovy JL, Taverna M
Lab. Chip - 8(2) :294-301 - DOI:10.1039/b715238h - 2008
A microreactor for proteinase K (PK)-mediated protein digestion was developed as a step towards the elaboration of a fully integrated microdevice for the detection of pathological prion protein (PrP). PK-grafted magnetic beads were immobilized inside a polydimethylsiloxane (PDMS) microchannel using a longitudinal magnetic field parallel to the flow direction and a magnetic field gradient, thereby forming a matrix for enzymatic digestion. This self-organization provided uniform pore sizes, a low flow resistance and a strong reaction efficiency due to a very thin diffusion layer. The microreactor's performance was first evaluated using a model substrate, succinyl-ala-ala-ala-paranitroanilide (SAAAP). Reaction kinetics were typically accelerated a hundred-fold as compared to conventional batch reactions. Reproducibility was around 98% for on-chip experiments. This microsystem was then applied to the digestion of prion protein from brain tissues. Controlled proteolysis could be obtained by varying the on-chip flow rate, while a complete proteolysis of normal protein was achieved in only three minutes. Extracts from normal and pathological brain homogenates were finally compared and strong discrimination between normal and pathological samples was demonstrated.
Microfluidic droplet-based liquid-liquid extraction
Laboratoire Microfluidique, MEMS et nanostructures - P. Mary, V. Studer, P. Tabeling
Anal. Chem. - 80(8) :2680-7 - DOI:10.1021/ac800088s - 2008
We study microfluidic systems in which mass exchanges take place between moving water droplets, formed on-chip, and an external phase (octanol). Here, no chemical reaction takes place, and the mass exchanges are driven by a contrast in chemical potential between the dispersed and continuous phases. We analyze the case where the microfluidic droplets, occupying the entire width of the channel, extract a solutefluoresceinfrom the external phase (extraction) and the opposite case, where droplets reject a soluterhodamineinto the external phase (purification). Four flow configurations are investigated, based on straight or zigzag microchannels. Additionally to the experimental work, we performed two-dimensional numerical simulations. In the experiments, we analyze the influence of different parameters on the process (channel dimensions, fluid viscosities, flow rates, drop size, droplet spacing, ...). Several regimes are singled out. In agreement with the mass transfer theory of Young et al. (Young, W.; Pumir, A.; Pomeau, Y. Phys. Fluids A 1989, 1, 462), we find that, after a short transient, the amount of matter transferred across the droplet interface grows as the square root of time and the time it takes for the transfer process to be completed decreases as Pe-2/3, where Pe is the Peclet number based on droplet velocity and radius. The numerical simulation is found in excellent consistency with the experiment. In practice, the transfer time ranges between a fraction and a few seconds, which is much faster than conventional systems.
Nanofluidics in the Debye layer at hydrophilic and hydrophobic surfaces
Laboratoire Microfluidique, MEMS et nanostructures - C.I Bouzigues, P. Tabeling, L. Bocquet
Phys. Rev. Lett. - 101(11) :114503 - DOI:10.1103/PhysRevLett.101.114503 - 2008
By using evanescent waves, we study equilibrium and dynamical properties of liquid-solid interfaces in the Debye layer for hydrophilic and hydrophobic surfaces. We measure velocity profiles and nanotracer concentration and diffusion profiles between 20 and 300 nm from the walls in pressure-driven and electro-osmotic flows. We extract electrostatic and zeta potentials and determine hydrodynamic slip lengths with 10 nm accuracy. The spectacular amplification of the zeta potential resulting from hydrodynamic slippage allows us to clarify for the first time the dynamic origin of the zeta potential.
Theoretical study of sequence-dependent nanopore unzipping of DNA
Laboratoire Nanobiophysiques - U. Bockelmann and V. Viasnoff
Biophys. J. - 94(7) :2716–24 - DOI:10.1529/biophysj.107.111732 - 2008
We theoretically investigate the unzipping of DNA electrically driven through a nanometer-size pore. Taking the DNA base sequence explicitly into account, the unpairing and translocation process is described by a biased random walk in a one-dimensional energy landscape determined by the sequential basepair opening. Distributions of translocation times are numerically calculated as a function of applied voltage and temperature. We show that varying these two parameters changes the dynamics from a predominantly diffusive behavior to a dynamics governed by jumps over local energy barriers. The work suggests experimentally studying sequence effects, by comparing the average value and standard deviation of the statistical distribution of translocation times.
Interference and crosstalk in double optical tweezers using a single laser source
Laboratoire Nanobiophysiques - P. Mangeol and U. Bockelmann
Rev Sci Instrum - 79(8) :083103 - DOI:10.1063/1.2957652 - 2008
Experimental studies of single molecule mechanics require high force sensitivity and low drift, which can be achieved with optical tweezers. We built an optical tweezer setup for force measurements in a two bead assay. A cw infrared laser beam is split by polarization and focused by a high numerical aperture objective to create two traps. The same laser is used to form both traps and to measure the force by back focal plane interferometry. We show that although the two beams entering the microscope are designed to exhibit orthogonal polarization, interference and a significant parasitic force signal occur. Comparing the experimental results with a ray optics model, we show that the interference patterns are caused by the rotation of polarization on microscope lens surfaces and slides. The model qualitatively describes the pattern and the dependence of the parasitic force signal on the experimental parameters. We present two different approaches to experimentally reduce the crosstalk, namely, polarization rectification and frequency shifting.
A caged retinoic acid for one- and two-photon excitation in zebrafish embryos
Laboratoire Physique des biomolécules - P. Neveu, I. Aujard, C. Benbrahim, T. Le Saux, J.-F. Allemand, S. Vriz, D. Bensimon and L. Jullien
Angew Chem Int Ed Engl. - 47(20) :3744–46 - DOI:10.1002/anie.200800037 - 2008
Specific Wetting Probed With Biomimetic Droplets
Laboratoire Pôle Microfluidique - J. Fattaccioli, J. Baudry, F. Brochard-Wyart, N. Henry and J. Bibette
Soft Matter - 4(12) :2334-40 - DOI:10.1039/B806635C - 2008
We have produced emulsion droplets of controlled size and composition coated by ligands, and studied the adhesion of these drops on a solid substrate coated by receptors and polymers. Using transmission, RICM and fluorescence microscopy we assess the size, contact angle and ligand density for each drop. We first show that non-specific interactions significantly enhance the proteins density within the adhesive patch. Then we show that binding within the patch is partially inhibited in good agreement with the hypothesis of an absence of translational diffusion. We confirm that the density of specific bonds sets the adhesive energy and therefore the final contact angle, and finally show that specific binding in our system is always associated with the existence of a positive line tension, which linearly increases with the density of receptors. These experiments describe a new scenario for specific wetting which raises the importance of the coupling between non-specific interactions and specific binding.
In situ quantitative measurement of concentration profiles in a microreactor with submicron resolution using multiplex CARS microscopy
Laboratoire Procédés, Plasmas, Microsystèmes - Dawn Schafer, Jeff A. Squier, Jan van Maarseveen, Daniel Bon, Mischa Bonn, Michiel Mu¨ller
JACS - 130(35) :11592-3 - DOI:10.1021/ja804158n - 2008
In situ quantitative imaging of concentration profiles of reactants and products inside a microfluidic reactor is achieved, with submicron spatial resolution with mM sensitivity and on ms time scales, for a given position. The label-free approach relies on quantitative vibrational spectroscopy, using Coherent Anti-Stokes Raman scattering microscopy in a spectrally resolved fashion, and is demonstrated on an elementary acid-base reaction.
A simple and universal tool to remove on-line impurities in mono- or two-dimensional liquid chromatography–mass spectrometry analysis
Laboratoire Spectrométrie de masse biologique et protéomique - Hesse A-M, Marcelo P, Rossier J and Vinh J.
J. Chrom. A - 1189(1-2) :175-82 - DOI:10.1016/j.chroma.2007.12.060 - 2008
Several recurrent problems have always hindered mono-dimensional liquid chromatography-mass spectrometry proteomic analyses. Polymer contamination is a major problem because polymers could co-elute with compounds of interest (peptides). In this case spectral suppression degrades dynamic range and sensitivity. Polyethylene glycol derivatives count among the major contaminants. They are targeted in this work. They are eluted at 35-40% acetonitrile from C18 phase in every single reversed-phase run. Moreover, they are also observed in two-dimensional liquid chromatography in every salt fraction. A simple and robust method is presented here for rapid and efficient on-line removal of these impurities using self-regenerating purification microdevices.
Determination of nanoparticle diffusion coefficients by Taylor dispersion analysis using a capillary electrophoresis instrument
Laboratoire Synthèse, Electrochimie, Imagerie et Systèmes Analytiques... - d'Orlye F, Varenne A, Gareil P.
J. Chrom. A - 1204(2) :226-32 - DOI:10.1016/j.chroma.2008.08.008 - 2008
The collective diffusion coefficient D(C) of diluted suspensions of positively charged iron oxide maghemite particles was experimentally investigated using a capillary electrophoresis instrument on the grounds of Taylor dispersion theory. Conditions for this approach to be applicable to nanoparticles of mean solid diameter below 10nm were set in this work, enabling precisions on D(C) determination of less than 2% relative standard deviation (RSD). Significantly different D(C) values were thus measured for particle populations differing in solid number mean diameter by only 2 nm. The obtained values were compared to the z-average diffusion coefficient derived from dynamic light scattering (DLS) experiments and used for the calculation of the Stokes radius. The measured diffusion coefficients appeared to be dependent on particle volume fraction and electrolyte ionic strength. These observations were eventually discussed in terms of particle interactions.

410 publications.