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On-Chip Quantitative Measurement of Mechanical Stresses During Cell Migration with Emulsion Droplets
Laboratoire Pôle Microfluidique - D. Molino, S. Quignard, C. Gruget, F. Pincet, Y. Chen, M. Piel & J. Fattaccioli
Scientific Reports - 6 29113 - DOI: 10.1038/srep29113 - 2019
The ability of immune cells to migrate within narrow and crowded spaces is a critical feature involved in various physiological processes from immune response to metastasis. Several in-vitro techniques have been developed so far to study the behaviour of migrating cells, the most recent being based on the fabrication of microchannels within which cells move. To address the question of the mechanical stress a cell is able to produce during the encounter of an obstacle while migrating, we developed a hybrid microchip made of parallel PDMS channels in which oil droplets are sparsely distributed and serve as deformable obstacles. We thus show that cells strongly deform droplets while passing them. Then, we show that the microdevice can be used to study the influence of drugs on migration at the population level. Finally, we describe a quantitative analysis method of the droplet deformation that allows measuring in real-time the mechanical stress exerted by a single cell. The method presented herein thus constitutes a powerful analytical tool for cell migration studies under confinement.
Nano-on-Micro Fibrous Extracellular Matrices for Scalable Expansion of Human Es/Ips Cells
Laboratoire Pôle Microfluidique - L. Liu, K.-i. Kamei, M. Yoshioka, M. Nakajima, J. Li, N. Fujimoto, S. Terada, Y. Tokunaga, Y. Koyama, H. Sato, K. Hasegawa, N. Nakatsuji and Y. Chen
Biomaterials - 124 47-54 - DOI: 10.1016/j.biomaterials.2017.01.039 - 2019
Human pluripotent stem cells (hPSCs) hold great potential for industrial and clinical applications. Clinical-grade scaffolds and high-quality hPSCs are required for cell expansion as well as easy handling and manipulation of the products. Current hPSC culture methods do not fulfill these requirements because of a lack of proper extracellular matrices (ECMs) and cell culture wares. We developed a layered nano-on-micro fibrous cellular matrix mimicking ECM, named "fiber-on-fiber (FF)" matrix, which enables easy handling and manipulation of cultured cells. While non-woven sheets of cellulose and polyglycolic acid were used as a microfiber layer facilitating mechanical stability, electrospun gelatin nanofibers were crosslinked on the microfiber layer, generating a mesh structure with connected nanofibers facilitating cell adhesion and growth. Our results showed that the FF matrix supports effective hPSC culture with maintenance of their pluripotency and normal chromosomes over two months, as well as effective scaled-up expansion, with fold increases of 54.1 ± 15.6 and 40.4 ± 8.4 in cell number per week for H1 human embryonic stem cells and 253G1 human induced pluripotent stem cells, respectively. This simple approach to mimick the ECM may have important implications after further optimization to generate lineage-specific products.
Patterned parylene C for cell adhesion, spreading and alignment studies
Laboratoire Pôle Microfluidique - Xiaolong Tu, JinWei, BinWang, Yadong Tanga, Jian Shi, Yong Chen
ELSEVIER - 175 56–60 - - 2017
Parylene C iswidely used for insulating coating of implantablemedical devices due to its unique properties of biocompatibility
and biostability.However, the interaction between cells and parylene C is limited with as-deposited
samples. A plasma treatment of the sample can significantly improve the cell adhesion on parylene C. To further
improve the cell adhesion as well as other cell functions, we patterned parylene C into pillars and stripes by soft
lithography and chemical vapor deposition. Such amethod is flexible to produce parylene C patterns on different
surfaces. The fabricated substrates have been tested to culture Hela cells, showing much improved adhesion on
pillar arrays of parylene C. Our results also showed a strong effect of contact guidance for cells on stripes of
parylene C, thereby proving the relevance of the method for cell adhesion, spreading and alignment studies
Photosensitive Polyamines for High-Performance Photocontrol of DNA Higher-Order Structure Venancio-Marques
Laboratoire Pôle Microfluidique - Venancio-Marques, Anna, Bergen Anna, Rossi-Gendron Caroline, Rudiuk Sergii, and Baigl Damien
American Chemical Society Nano (ACS Nano) - Volume 8 (4) 3654–3663 - DOI: 10.1021/nn500266b - 2014
Polyamines are small, ubiquitous, positively charged molecules that play an essential role in numerous biological processes such as DNA packaging, gene regulation, neuron activity, and cell proliferation. Here, we synthesize the first series of photosensitive polyamines (PPAs) and demonstrate their ability to photoreversibly control nanoscale DNA higher-order structure with high efficiency. We show with fluorescence microscopy imaging that the efficiency of the PPAs as DNA-compacting agents is directly correlated to their molecular charge. Micromolar concentration of the most efficient molecule described here, a PPA containing three charges at neutral pH, compacts DNA molecules from a few kilobase pairs to a few hundred kilobase pairs, while subsequent 3 min UV illuminations at 365 nm triggers complete unfolding of DNA molecules. Additional application of blue light (440 nm for 3 min) induces the refolding of DNA into the compact state. Atomic force microscopy reveals that the compaction involves a global folding of the whole DNA molecule, whereas UV-induced unfolding is a modification initiated from the periphery of the compacted DNA, resulting in the occurrence of intermediate flower-like structures prior to the fully unfolded state.

Keywords: polyamines; DNA compaction; photocontrol; DNA; AFM; light
Dynamic photocontrol of the coffee-ring effect with optically-tunable particle stickiness
Laboratoire Pôle Microfluidique - Dr. Manos Anyfantakis and Prof. Damien Baigl
Angew Chem Int Ed Engl. - Volume 53, Issue 51 14077–14081 - DOI: 10.1002/anie.201406903 - 2014
When a colloidal drop dries on a surface, most of the particles accumulate at the drop periphery, yielding a characteristic ring-shaped pattern. This so-called coffee-ring effect (CRE) is observed in any pinned evaporating drop containing non-volatile solutes. Here, the CRE is dynamically controlled for the first time by using light, and an unprecedented reconfigurability of the deposit profile is demonstrated. This is achieved through a new mechanism where particle stickiness is optically tuned on demand, thus offering reliable modulation of the deposition pattern. The system consists of anionic nanoparticles and photosensitive cationic surfactants dispersed in water. It is shown that light-dependent modulation of surfactant–particle interactions dictates particle attraction and trapping at the liquid–gas interface, which allows us to direct particle deposition into a wide range of patterns from rings to homogeneous disks. Patterning from single drops is photoreversible upon changing the wavelength whereas spatial control in multiple drop arrays is achieved using a photomask.
Microfluidic device with integrated microfilter of conical-shaped holes for high efficiency and high purity capture of circulating tumor cells
Laboratoire Pôle Microfluidique - Yadong Tang, Jian Shi, Sisi Li, Li Wang, Yvon E. Cayre and Yong Chen
Scientific Reports - 4 (n°6052) - DOI:10.1038/srep06052 - 2013
Capture of circulating tumor cells (CTCs) from peripheral blood of cancer patients has major implications for metastatic detection and therapy analyses. Here we demonstrated a microfluidic device for high efficiency and high purity capture of CTCs. The key novelty of this approach lies on the integration of a microfilter with conical-shaped holes and a micro-injector with cross-flow components for size dependent capture of tumor cells without significant retention of non-tumor cells. Under conditions of constant flow rate, tumor cells spiked into phosphate buffered saline could be recovered and then cultured for further analyses. When tumor cells were spiked in blood of healthy donors, they could also be recovered at high efficiency and high clearance efficiency of white blood cells. When the same device was used for clinical validation, CTCs could be detected in blood samples of cancer patients but not in that of healthy donors. Finally, the capture efficiency of tumor cells is cell-type dependent but the hole size of the filter should be more closely correlated to the nuclei size of the tumor cells. Together with the advantage of easy operation, low-cost and high potential of integration, this approach offers unprecedented opportunities for metastatic detection and cancer treatment monitoring.
UV-Induced Bursting of Cell-Sized Multicomponent Lipid Vesicles in a Photosensitive Surfactant Solution
Laboratoire Pôle Microfluidique - A. Diguet, M. Yanagisawa, Y. J. Liu, E. Brun, S. Abadie, S. Rudiuk and D. Baigl
JACS - 134(10) :4898-904 - DOI:10.1021/ja211664f - 2012
We study the behavior of multicomponent giant unilamellar vesicles (GUVs) in the presence of AzoTAB, a photosensitive surfactant. GUVs are made of an equimolar ratio of dioleoylphosphatidylcholine (DOPC) and dipalmitoylphosphatidylcholine (DPPC) and various amounts of cholesterol (Chol), where the lipid membrane shows a phase separation into a DPPC-rich liquid-ordered (Lo) phase and a DOPC-rich liquid-disordered (Ld) phase. We find that UV illumination at 365 nm for 1 s induces the bursting of a significant fraction of the GUV population. The percentage of UV-induced disrupted vesicles, called bursting rate (Yburst), increases with an increase in [AzoTAB] and depends on [Chol] in a non-monotonous manner. Yburst decreases when [Chol] increases from 0 to 10 mol % and then increases with a further increase in [Chol], which can be correlated with the phase composition of the membrane. We show that Yburst increases with the appearance of solid domains ([Chol] = 0) or with an increase in area fraction of Lo phase (with increasing [Chol] = 10 mol %). Under our conditions (UV illumination at 365 nm for 1 s), maximal bursting efficiency (Yburst = 53%) is obtained for [AzoTAB] = 1 mM and [Chol] = 40 mol %. Finally, by restricting the illumination area, we demonstrate the first selective UV-induced bursting of individual target GUVs. These results show a new method to probe biomembrane mechanical properties using light as well as pave the way for novel strategies of light-induced drug delivery.
Integrated and Diffusion-Based Micro-Injectors for Open Access Cell Assays
Laboratoire Pôle Microfluidique - X. Li, L. Liu, L. Wang, K. Kamei, Q. H. Yuan, F. Zhang, J. Shi, A. Kusumi, M. Xie, Z. J. Zhao and Y. Chen
Lab. Chip - 11 :2612-7 - DOI:10.1039/c1lc20258h - 2011
Currently, most microfluidic devices are fabricated with embedded micro-channels and other elements in a close form with outward connections. Although much functionality has been demonstrated and a large number of applications have been developed, they are not easy for routine operation in biology laboratories where most in vitro cell processing still relies on the use of culture dishes, glass slides, multi-well plates, tubes, pipettes, etc. We report here an open access device which consists of an array of isolated micro-channels plated on a large culture surface, each of them having tiny nozzles for localized drug delivery. In a diffusion dominant regime, steady gradients of molecule concentration could be obtained and varied by changing the flow rate inside the micro-channels. As assay examples, cell staining and drug-induced cell apoptosis were demonstrated, showing fast cell responses in close proximity of the nozzles.
Photoreversible Fragmentation of a Liquid Interface for Micro-Droplet Generation by Light Actuation
Laboratoire Pôle Microfluidique - A. Diguet, H. Li, N. Queyriaux, Y. Chen and D. Baigl
Lab. Chip - 11 :2666-9 - DOI:10.1039/c1lc20328b - 2011
We describe a method to induce by light a reversible switch from a continuous two-phase laminar flow to a droplet generating regime, in microfluidic devices with a usual water-in-oil flow focusing geometry. It consists in adding a photosensitive surfactant to the aqueous phase to modulate using light the interfacial energy between flowing liquids and the microfluidic substrate. We show that UV irradiation induces liquid fragmentation into monodisperse water microdroplets and that many cycles of reversible and rapid switches (<2 s) between continuous laminar flows and stable droplet regimes can be realized. By spatially controlling the application of the light stimulus, we also demonstrate the first spatially resolved remote induction of droplet generation.
Pre-Dispositions and Epigenetic Inheritance in the Escherichia Coli Lactose Operon Bistable Switch
Laboratoire Pôle Microfluidique - L. Robert, G. Paul, Y. Chen, F. Taddei, D. Baigl and A. B. Lindner
Molecular Systems Biology - 6 :357 - DOI:10.1038/msb.2010.12 - 2010
The lactose operon regulation in Escherichia coli is a primary model of phenotypic switching, reminiscent of cell fate determination in higher organisms. Under conditions of bistability, an isogenic cell population partitions into two subpopulations, with the operon's genes turned on or remaining off. It is generally hypothesized that the final state of a cell depends solely on stochastic fluctuations of the network's protein concentrations, particularly on bursts of lactose permease expression. Nevertheless, the mechanisms underlying the cell switching decision are not fully understood. We designed a microfluidic system to follow the formation of a transiently bimodal population within growing microcolonies. The analysis of genealogy and cell history revealed the existence of pre-disposing factors for switching that are epigenetically inherited. Both the pre-induction expression stochasticity of the lactose operon repressor LacI and the cellular growth rate are predictive factors of the cell's response upon induction, with low LacI concentration and slow growth correlating with higher switching probability. Thus, stochasticity at the local level of the network and global physiology are synergistically involved in cell response determination.
Photomanipulation of a Droplet by the Chromocapillary Effect
Laboratoire Pôle Microfluidique - A. Diguet, R. M. Guillermic, N. Magome, A. Saint-Jalmes, Y. Chen, K. Yoshikawa and D. Baigl
Angew Chem Int Ed Engl. - 48(49) :9281-4 - DOI:10.1002/anie.200904868 - 2009
Ace of hearts: Liquid droplets can be manipulated in a controlled fashion along trajectories of any desired shape (such as a heart, see picture) by using light to create a wavelength-dependent interfacial tension gradient at a liquid/liquid interface. In this new phenomenon, the “chromocapillary effect”, an interfacial flow generates droplet motion in the direction opposite to the gradient.
Sequence-Independent and Reversible Photocontrol of Transcription/Expression Systems Using a Photosensitive Nucleic Acid Binder
Laboratoire Pôle Microfluidique - A. Estevez-Torres, C. Crozatier, A. Diguet, T. Hara, H. Saito, K. Yoshikawa and D. Baigl
Proc. Nat. Acad. Sci. USA - 106(30) :12219-23 - DOI:10.1073/pnas.0904382106 - 2009
To understand non-trivial biological functions, it is crucial to develop minimal synthetic models that capture their basic features. Here, we demonstrate a sequence-independent, reversible control of transcription and gene expression using a photosensitive nucleic acid binder (pNAB). By introducing a pNAB whose affinity for nucleic acids is tuned by light, in vitro RNA production, EGFP translation, and GFP expression (a set of reactions including both transcription and translation) were successfully inhibited in the dark and recovered after a short illumination at 365 nm. Our results indicate that the accessibility of the protein machinery to one or several nucleic acid binding sites can be efficiently regulated by changing the conformational/condensation state of the nucleic acid (DNA conformation or mRNA aggregation), thus regulating gene activity in an efficient, reversible, and sequence-independent manner. The possibility offered by our approach to use light to trigger various gene expression systems in a system-independent way opens interesting perspectives to study gene expression dynamics as well as to develop photocontrolled biotechnological procedures.
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.
Highly Parallel Mix-and-Match Fabrication of Nanopillar Arrays Integrated in Microfluidic Channels for Long DNA Molecule Separation
Laboratoire Pôle Microfluidique - J. Shi, A. P. Fang, L. Malaquin, A. Pepin, D. Decanini, J. L. Viovy and Y. Chen
Applied Physics Letters - 91(15) :153114 - DOI:153114 10.1063/1.2793616 - 2007
We report on a mix-and-match method based on a combination of soft UV nanoimprint lithography, contact optical lithography, and reactive-ion-etch techniques, which is applicable for high throughput manufacturing of nanostructure integrated microfluidic devices. We demonstrate the integration of high density and high aspect ratio nanopillars into microfluidic channels as electrophoresis sieving matrices. As a result, ? DNA and T4 DNA can be separated within a few minutes. By changing the pattern design, the device could be used for separation of other types of molecules.
Microcontact Printing of Living Bacteria Arrays with Cellular Resolution
Laboratoire Pôle Microfluidique - L. P. Xu, L. Robert, O. Y. Qi, F. Taddei, Y. Chen, A.B. Lindner, D. Baigl
Nano Lett. - 7(7) :2068-72 - DOI:10.1021/nl070983z - 2007
Arrays of living bacteria were printed on agarose substrate with cellular resolution using elastomeric stamps with a high aspect ratio generated by reverse in situ lithography (RISL). The printed bacteria reproduced the original stamp patterns with high fidelity and continued growing as in bulk culture. This methodology provides a simple route to any desired bacterial spatial 2D distribution and may be applied to screening as well as to studies of bacteria phenotypic variability, population dynamics, and ecosystem evolution.

15 publications.