A microfluidic platform integrating functional vascularized organoids-on-chip.

The development of vascular networks in microfluidic chips is crucial for the long-term culture of three-dimensional cell aggregates such as spheroids, organoids, tumoroids, or tissue explants. Despite rapid advancement in microvascular network systems and organoid technologies, vascularizing organoids-on-chips remains a challenge in tissue engineering. Most existing microfluidic devices poorly reflect the complexity of in vivo flows and require complex technical set-ups. Considering these constraints, we develop a platform to establish and monitor the formation of endothelial networks around mesenchymal and pancreatic islet spheroids, as well as blood vessel organoids generated from pluripotent stem cells, cultured for up to 30 days on-chip. We show that these networks establish functional connections with the endothelium-rich spheroids and vascular organoids, as they successfully provide intravascular perfusion to these structures. We find that organoid growth, maturation, and function are enhanced when cultured on-chip using our vascularization method. This microphysiological system represents a viable organ-on-chip model to vascularize diverse biological 3D tissues and sets the stage to establish organoid perfusions using advanced microfluidics.

Microfluidic device integrating a network of hyper-elastic valves for automated glucose stimulation and insulin secretion collection from a single pancreatic islet.

Advances in microphysiological systems have prompted the need for robust and reliable cell culture devices. While microfluidic technology has made significant progress, devices often lack user-friendliness and are not designed to be industrialized on a large scale. Pancreatic islets are often being studied using microfluidic platforms in which the monitoring of fluxes is generally very limited, especially because the integration of valves to direct the flow is difficult to achieve. Considering these constraints, we present a thermoplastic manufactured microfluidic chip with an automated control of fluxes for the stimulation and secretion collection of pancreatic islet. The islet was directed toward precise locations through passive hydrodynamic trapping and both dynamic glucose stimulation and insulin harvesting were done automatically via a network of large deformation valves, directing the reagents and the pancreatic islet toward different pathways. This device we developed enables monitoring of insulin secretion from a single islet and can be adapted for the study of a wide variety of biological tissues and secretomes.

A versatile and automated microfluidic platform for a quantitative magnetic bead based protocol: application to gluten detection.

A microfluidic platform for the integration of multi-step biological assays has been developed. The presented system is a unique instrument compatible with microfluidic chips for various applications based on bead manipulation. Two examples of microfluidic cartridges are presented here. The first one contains two rows of eight chambers (40 and 80 µL), six reagent inlets, eight testing solution (calibrators and samples) inlets and eight outlets to reproduce precisely each step of a biological assay. This configuration is versatile enough to integrate many different biological assays and save a lot of development time. The second architecture is dedicated to one specific protocol and is completely automated from the standard and sample dilutions to the optical detection. Linear dilutions have been integrated to prepare automatically a range of standard concentrations and outlets have been modified for integrated colorimetric detection. The technology uses pneumatically collapsible chambers to perform all the fluidic operations for a fully automated protocol such as volume calibrations, fluid transport, mixing, and washing steps. A programmable instrument with a software interface has been developed to adapt rapidly a protocol to this cartridge. As an example, these new microfluidic cartridges have been used to successfully perform an immunoassay for gluten detection in the dynamic range of 10-30 ppm with good sensitivity (2 ppm) and specificity.

Selective plane illumination microscope dedicated to volumetric imaging in microfluidic chambers.

In this article, we are presenting an original selective plane illumination fluorescence microscope dedicated to image "Organ-on-chip"-like biostructures in microfluidic chips. In order to be able to morphologically analyze volumetric samples in development at the cellular scale inside microfluidic chambers, the setup presents a compromise between relatively large field of view (∼ 200 µm) and moderate resolution (∼ 5 µm). The microscope is based on a simple design, built around the chip and its microfluidic environment to allow 3D imaging inside the chip. In particular, the sample remains horizontally avoiding to disturb the fluidics phenomena. The experimental setup, its optical characterization and the first volumetric images are reported.

Fast and continuous plasma extraction from whole human blood based on expanding cell-free layer devices.

This paper presents promising microfluidic devices designed for continuous and passive extraction of plasma from whole human blood. These designs are based on red cells lateral migration and the resulting cell-free layer locally expanded by geometric singularities such as an enlargement of the channel or a cavity adjacent to the channel. After an explanation of flow patterns, different tests are described that confirm the advantages of both proposed singularities, providing a 1.5 and 2X increase in extraction yield compared to a reference device, for 1:20 diluted blood at 100 microL/min. Devices have also been successively optimized, with extraction yields up to 17.8%, and biologically validated for plasma extraction, with no protein loss or denaturation, no hemolysis and with excellent cell purity. Finally, the dilution effect has been experimentally investigated.

Separation of Biological Particles in a Modular Platform of Cascaded Deterministic Lateral Displacement Modules.

Deterministic lateral displacement (DLD) has been extensively implemented in the last decade for size-based sample preparation, owing to its high separation performances for a wide range of particle dimensions. However, separating particles from 1 µm to 10 µm in one single DLD device is challenging because of the required diversity of pillar dimensions and inherent fabrication issues. This paper presents an alternative approach to achieve the extraction of E. coli bacteria from blood samples spiked with prostate cancer cells. Our approach consists in cascading individual DLD devices in a single automated platform, using flexible chambers that successively collect and inject the sample between each DLD stage without any external sample manipulation. Operating DLD separations independently enables to maximize the sorting efficiency at each step, without any disturbance from downstream stages. The proposed two-step automated protocol is applied to the separation of three types of components (bacteria, blood particles and cancer cells), with a depletion yield of 100% for cancer cells and 93% for red blood cells. This cascaded approach is presented for the first time with two DLD modules and is upscalable to improve the dynamic range of currently available DLD devices.

Coplanar electrowetting-induced stirring as a tool to manipulate biological samples in lubricated digital microfluidics. Impact of ambient phase on drop internal flow pattern.

Oscillating electrowetting on dielectrics (EWOD) with coplanar electrodes is investigated in this paper as a way to provide efficient stirring within a drop with biological content. A supporting model inspired from Ko et al. [Appl. Phys. Lett. 94, 194102 (2009)] is proposed allowing to interpret oscillating EWOD-induced drop internal flow as the result of a current streaming along the drop surface deformed by capillary waves. Current streaming behaves essentially as a surface flow generator and the momentum it sustains within the (viscous) drop is even more significant as the surface to volume ratio is small. With the circular electrode pair considered in this paper, oscillating EWOD sustains toroidal vortical flows when the experiments are conducted with aqueous drops in air as ambient phase. But when oil is used as ambient phase, it is demonstrated that the presence of an electrode gap is responsible for a change in drop shape: a pinch-off at the electrode gap yields a peanut-shaped drop and a symmetry break-up of the EWOD-induced flow pattern. Viscosity of oil is also responsible for promoting an efficient damping of the capillary waves which populate the surface of the actuated drop. As a result, the capillary network switches from one standing wave to two superimposed traveling waves of different mechanical energy, provided that actuation frequency is large enough, for instance, as large as the one commonly used in electrowetting applications (f ∼ 500 Hz and beyond). Special emphasis is put on stirring of biological samples. As a typical application, it is demonstrated how beads or cell clusters can be focused under flow either at mid-height of the drop or near the wetting plane, depending on how the nature of the capillary waves is (standing or traveling), and therefore, depending on the actuation frequency (150 Hz-1 KHz).

A programmable and reconfigurable microfluidic chip.

This article reports an original concept enabling the rapid fabrication of continuous-flow microfluidic chips with a programmable and reconfigurable geometry. The concept is based on a digital microfluidic platform featuring an array of individually addressable electrodes. A selection of electrodes is switched on sequentially to create a de-ionized (DI) water finger specific pattern, while the surrounding medium consists of liquid-phase paraffin. The water displacement is induced by both electrowetting on dielectric and liquid dielectrophoresis phenomena. Once the targeted DI water pattern is obtained, the chip temperature is lowered by turning on an integrated thermoelectric cooler, forming channel structures made of solidified paraffin with edges delimitated by the DI water pattern. As a result, the chip can be used afterwards to conduct in-flow continuous microfluidic experiments. This process is resettable and reversible by heating up the chip to melt the paraffin and reconfigure the microchannel design on demand, offering the advantages of cost, adaptability, and robustness. This paper reports experimental results describing the overall concept, which is illustrated with typical and basic fluidic geometries.

Enrichment of nanoparticles and bacteria using electroless and manual actuation modes of a bypass nanofluidic device.

Current efforts in nanofluidics aimed at detecting scarce molecules or particles are focused mainly on the development of electrokinetic-based devices. However, these techniques require either integrated or external electrodes, and a potential drop applied across a carrier fluid. One challenge is to develop a new generation of electroless passive devices involving a simple technological process and packaging without embedded electrodes for micro- and nanoparticles enrichment with a view to applications in biology such as the detection of viral agents or cancers biomarkers. This paper presents an innovative technique for particles handling and enrichment based exclusively on a pressure-driven silicon bypass nanofluidic device. The device is fabricated by standard silicon micro-nanofabrication technology. The concentration operation was demonstrated and quantified according to two different actuation modes, which can also be combined to enhance the concentration factor further. The first, "symmetrical" mode involves a symmetric cross-flow effect that concentrates nanoparticles in a very small volume in a very local point of the device. The second mode, "asymmetrical" mode advantageously generates a streaming potential, giving rise to an Electroless Electropreconcentration (EL-EP). The concentration process can be maintained for several hours and concentration factors as high as ~200 have been obtained when both symmetrical and asymmetrical modes are coupled. Proof of concept for concentrating E. coli bacteria by the manual actuation of the EL-EP device is also demonstrated in this paper. Experiments demonstrate more than a 50-fold increase in the concentration of E. coli bacteria in only ~40 s.

Macro to microfluidics system for biological environmental monitoring.

Biological environmental monitoring (BEM) is a growing field of research which challenges both microfluidics and system automation. The aim is to develop a transportable system with analysis throughput which satisfies the requirements: (i) fully autonomous, (ii) complete protocol integration from sample collection to final analysis, (iii) detection of diluted molecules or biological species in a large real life environmental sample volume, (iv) robustness and (v) flexibility and versatility. This paper discusses all these specifications in order to define an original fluidic architecture based on three connected modules, a sampling module, a sample preparation module and a detection module. The sample preparation module highly concentrates on the pathogens present in a few mL samples of complex and unknown solutions and purifies the pathogens' nucleic acids into a few µL of a controlled buffer. To do so, a two-step concentration protocol based on magnetic beads is automated in a reusable macro-to-micro fluidic system. The detection module is a PCR based miniaturized platform using digital microfluidics, where reactions are performed in 64 nL droplets handled by electrowetting on dielectric (EWOD) actuation. The design and manufacture of the two modules are reported as well as their respective performances. To demonstrate the integration of the complete protocol in the same system, first results of pathogen detection are shown.

Ionic liquid droplet as e-microreactor.

A powerful approach combining a droplet-based, open digital microfluidic lab-on-a-chip using task-specific ionic liquids as soluble supports to perform solution-phase synthesis is reported as a new tool for chemical applications. The negligible volatility of ionic liquids enables their use as stable droplet reactors on a chip surface under air. The concept was validated with different ionic liquids and with a multicomponent reaction. Indeed, we showed that different ionic liquids can be moved by electrowetting on dielectric (EWOD), and their displacement was compared with aqueous solutions. Furthermore, we showed that mixing ionic liquids droplets, each containing a different reagent, in "open" systems is an efficient way of carrying supported organic synthesis. This was applied to Grieco's tetrahydroquinolines synthesis with different reagents. Analysis of the final product was performed off-line and on-line, and the results were compared with those obtained in a conventional reaction flask. This technology opens the way to easy synthesis of minute amounts of compounds ad libitum without the use of complex, expensive, and bulky robots and allows complete automation of the process for embedded chemistry in a portable device. It offers several advantages, including simplicity of use, flexibility, and scalability, and appears to be complementary to conventional microfluidic lab-on-a-chip devices usually based on continuous-flow in microchannels.