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Chromatin immunoprecipitation in microfluidic droplets: towards fast and cheap analyses
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.
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
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.
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.
Optimizing Hyperuniformity in Self-Assembled Bidisperse Emulsions
Joshua Ricouvier, Romain Pierrat, Rémi Carminati, Patrick Tabeling, and Pavel Yazhgur
Phys. Rev. Lett. - 119 208001 - doi.org/10.1103/PhysRevLett. - 2017
We study long range density fluctuations (hyperuniformity) in two-dimensional jammed packings of bidisperse droplets. Taking advantage of microfluidics, we systematically span a large range of size and concentration ratios of the two droplet populations. We identify various defects increasing long range density fluctuations mainly due to organization of local particle environment. By choosing an appropriate bidispersity, we fabricate materials with a high level of hyperuniformity. Interesting transparency properties of these optimized materials are established based on numerical simulations.
Roughness of oxide glass subcritical fracture surfaces
Gael Pallares , Frederic Lechenault, Matthieu George, Elisabeth Bouchaud, Cédric Ottina, Cindy L. Rountree, Matteo Ciccotti ,
Phys. Chem. - 101 (3) 1279-1288 - DOI : 10.1111/jace.15262 - 2017
An original setup combining a very stable loading stage, an atomic force microscope and an environmental chamber, allows to obtain very stable sub-critical fracture propagation in oxide glasses under controlled environment, and subsequently to finely characterize the nanometric roughness properties of the crack surfaces. The analysis of the surface roughness is conducted both in terms of the classical root mean square roughness to compare with the literature, and in terms of more physically adequate indicators related to the self-affine nature of the fracture surfaces. Due to the comparable nanometric scale of the surface roughness, the AFM tip size and the instrumental noise, a special care is devoted to the statistical evaluation of the metrologic properties. The 2 roughness amplitude of several oxide glasses was shown to decrease as a function of the stress intensity factor, to be quite insensitive to the relative humidity and to increase with the degree of heterogeneity of the glass. The results are discussed in terms of several modeling arguments concerning the coupling between crack propagation, material's heterogeneity, crack tip plastic deformation and water diffusion at the crack tip. A synthetic new model is presented combining the predictions of a model by Wiederhorn et al. [1] on the effect of the material's heterogeneity on the crack tip stresses with the self-affine nature of the fracture surfaces.
Paper-based RNA detection and multiplexed analysis for Ebola virus diagnostics
Laura Magro, Béatrice Jacquelin, Camille Escadafal, Pierre Garneret, Aurélia Kwasiborski, Jean-Claude Manuguerra, Fabrice Monti, Anavaj Sakuntabhai, Jessica Vanhomwegen, Pierre Lafaye & Patrick Tabeling
Scientific Reports - 1347 (2017) - https://doi.org/10.1038/s41598-017-00758-9 - 2017
The most performing techniques enabling early diagnosis of infectious diseases rely on nucleic acid detection. Today, because of their high technicality and cost, nucleic acid amplification tests (NAAT) are of benefit only to a small fraction of developing countries population. By reducing costs, simplifying procedures and enabling multiplexing, paper microfluidics has the potential to considerably facilitate their accessibility. However, most of the studies performed in this area have not quit the lab. This letter brings NAAT on paper closer to the field, by using clinical samples and operating in a resource-limited setting. We first performed isothermal reverse transcription and Recombinase Polymerase Amplification (RT-RPA) of synthetic Ribonucleic Acid (RNA) of Ebola virus using paper microfluidics devices. We further applied this method in Guinea to detect the presence of Ebola virus in human sample RNA extracts, with minimal facilities (carry-on detection device and freeze-dried reagents on paper). RT-RPA results were available in few minutes and demonstrate a sensitivity of 90.0% compared to the gold-standard RT-PCR on a set of 43 patient samples. Furthermore, the realization of a nine-spot multilayered device achieving the parallel detection of three distinct RNA sequences opens a route toward the detection of multiple viral strains or pathogens.
In situ targeted activation of an anticancer agent using ultrasound-triggered release of composite droplets
Bezagu M, Clarhaut J, Renoux B, Monti F, Tanter M, Tabeling P, Cossy J, Couture O, Papot S, Arseniyadis S.
Eur J Med Chem. - 42 44014 - doi: 10.1016/j.ejmech.2017.03.057 - 2017
The efficiency of a drug is usually highly dependent on the way it is administered or delivered. As such, targeted-therapy, which requires conceiving drug-delivery vehicles that will change their state from a relatively stable structure with a very slow leak-rate to an unstable structure with a fast release, clearly improves the pharmacokinetics, the absorption, the distribution, the metabolism and the therapeutic index of a given drug. In this context, we have developed a particularly effective double stimuli-responsive drug-delivery method allowing an ultrasound-induced release of a monomethylauristatin E-glucuronide prodrug and its subsequent activation by a β-glucuronidase. This led to an increase of cytotoxicity of about 80% on cancer cells.
Microfluidic step-emulsification in axisymmetric geometry
Chakraborty, J. Ricouvier, P. Yazhgur, P. Tabelingb and A. M. Leshansky
Lab. Chip - 17 3609-3620 - DOI: 10.1039/C7LC00755H - 2017
Biphasic step-emulsification (Z. Li et al., Lab Chip, 2015, 15, 1023) is a promising microfluidic technique for high-throughput production of μm and sub-μm highly monodisperse droplets. The step-emulsifier consists of a shallow (Hele-Shaw) microchannel operating with two co-flowing immiscible liquids and an abrupt expansion (i.e., step) to a deep and wide reservoir. Under certain conditions the confined stream of the disperse phase, engulfed by the co-flowing continuous phase, breaks into small highly monodisperse droplets at the step. Theoretical investigation of the corresponding hydrodynamics is complicated due to the complex geometry of the planar device, calling for numerical approaches. However, direct numerical simulations of the three dimensional surface-tension-dominated biphasic flows in confined geometries are computationally expensive. In the present paper we study a model problem of axisymmetric step-emulsification. This setup consists of a stable core-annular biphasic flow in a cylindrical capillary tube connected co-axially to a reservoir tube of a larger diameter through a sudden expansion mimicking the edge of the planar step-emulsifier. We demonstrate that the axisymmetric setup exhibits similar regimes of droplet generation to the planar device. A detailed parametric study of the underlying hydrodynamics is feasible via inexpensive (two dimensional) simulations owing to the axial symmetry. The phase diagram quantifying the different regimes of droplet generation in terms of governing dimensionless parameters is presented. We show that in qualitative agreement with experiments in planar devices, the size of the droplets generated in the step-emulsification regime is independent of the capillary number and almost insensitive to the viscosity ratio. These findings confirm that the step-emulsification regime is solely controlled by surface tension. The numerical predictions are in excellent agreement with in-house experiments with the axisymmetric step-emulsifier.
Paper Microfluidics for Nucleic Acids Amplification Testing (NAAT) of Infectious Diseases
Magro L1, Escadafal C, Garneret P, Jacquelin B, Kwasiborski A, Manuguerra JC, Monti F, Sakuntabhai A, Vanhomwegen J, Lafaye P, Tabeling P.
Lab. Chip - 17(14) 2347-2371 - doi: 10.1039/c7lc00013h. - 2017
The diagnosis of infectious diseases is entering a new and interesting phase. Technologies based on paper microfluidics, coupled to developments in isothermal amplification of Nucleic Acids (NAs) raise opportunities for bringing the methods of molecular biology in the field, in a low setting environment. A lot of work has been performed in the domain over the last few years and the landscape of contributions is rich and diverse. Most often, the level of sample preparation differs, along with the sample nature, the amplification and detection methods, and the design of the device, among other features. In this review, we attempt to offer a structured description of the state of the art. The domain is not mature and there exist bottlenecks that hamper the realization of Nucleic Acid Amplification Tests (NAATs) complying with the constraints of the field in low and middle income countries. In this domain however, the pace of progress is impressively fast. This review is written for a broad Lab on a Chip audience.

410 publications.