Improved human islets' viability and functionality with mesenchymal stem cells and arg-gly-asp tripeptides supplementation of alginate micro-encapsulated islets in vitro.

INTRODUCTION: The extension of islet transplantation to a wider number of type 1 diabetes patients is compromised by severe adverse events related to the immunosuppressant therapy required for allogenic islet transplantation. In this context, microencapsulation offers the prospects of immunosuppressive-free therapy by physically isolating islets from the immune system. However, current biomaterials need to be optimized to: improve biocompatibility, guaranty the maintenance of graft viability and functionality, and prevent fibrosis overgrowth around the capsule in vivo. Accumulating evidence suggest that mesenchymal stem cells (MSCs) and anchor points consisting of tripeptides arg-gly-asp (RGD) have cytoprotective effects on pancreatic islets. Here, we investigated the effect of supplementing reference M-rich alginate microcapsules with MSCs and RGD-G rich alginate on bioprocessing as well as on human pancreatic islets viability and functionality. METHODS: We characterized the microcapsules components, and then for the new microcapsule composite product: we analyzed the empty capsules biocompatibility and then investigated the benefits of MSCs and RGD-G rich alginate on viability and functionality on the encapsulated human pancreatic islets in vitro. We performed viability tests by confocal microscopy and glucose stimulated insulin secretion (GSIS) test in vitro to assess the functionality of naked and encapsulated islets. RESULTS: Encapsulation in reference M-rich alginate capsules induced a reduction in viability and functionality compared to naked islets. This side-effect of encapsulation was in part counteracted by the presence of MSCs but the restoration was complete with the combination of both MSCs and the RGD-G rich alginate. CONCLUSIONS: The present findings show that bioprocessing a favorable composite environment inside the M-rich alginate capsule with both MSCs and RGD-G rich alginate improves human islets survival and functionality in vitro.

Anticipating Cutoff Diameters in Deterministic Lateral Displacement (DLD) Microfluidic Devices for an Optimized Particle Separation.

Deterministic lateral displacement (DLD) devices enable to separate nanometer to micrometer-sized particles around a cutoff diameter, during their transport through a microfluidic channel with slanted rows of pillars. In order to design appropriate DLD geometries for specific separation sizes, robust models are required to anticipate the value of the cutoff diameter. So far, the proposed models result in a single cutoff diameter for a given DLD geometry. This paper shows that the cutoff diameter actually varies along the DLD channel, especially in narrow pillar arrays. Experimental and numerical results reveal that the variation of the cutoff diameter is induced by boundary effects at the channel side walls, called the wall effect. The wall effect generates unexpected particle trajectories that may compromise the separation efficiency. In order to anticipate the wall effect when designing DLD devices, a predictive model is proposed in this work and has been validated experimentally. In addition to the usual geometrical parameters, a new parameter, the number of pillars in the channel cross dimension, is considered in this model to investigate its influence on the particle trajectories.

Isolation of Extracellular Vesicles from Cell Culture and Blood Through Nano-Targeted DLD Microfluidic Device.

Extracellular vesicles (EVs) are secreted by cells and found in biological fluids such as blood, with concentration correlated with oncogenic signals, making them attractive biomarkers for liquid biopsy. The current gold-standard method for EVs isolation requires an ultracentrifugation (UC) step among others. The cost and complexity of this technique are forbiddingly high for many researchers, as well as for routine use in biological laboratories and hospitals. This chapter reports on a simple microfluidic method for EVs isolation, based on a microfluidic size sorting technique named Deterministic Lateral Displacement (DLD). With the design of micrometric DLD array, we demonstrated the potential of our DLD devices for the isolation of nano-biological objects such as EVs, with main population size distribution consistent with UC technique.

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.

Purification of complex samples: Implementation of a modular and reconfigurable droplet-based microfluidic platform with cascaded deterministic lateral displacement separation modules.

Particle separation in microfluidic devices is a common problematic for sample preparation in biology. Deterministic lateral displacement (DLD) is efficiently implemented as a size-based fractionation technique to separate two populations of particles around a specific size. However, real biological samples contain components of many different sizes and a single DLD separation step is not sufficient to purify these complex samples. When connecting several DLD modules in series, pressure balancing at the DLD outlets of each step becomes critical to ensure an optimal separation efficiency. A generic microfluidic platform is presented in this paper to optimize pressure balancing, when DLD separation is connected either to another DLD module or to a different microfluidic function. This is made possible by generating droplets at T-junctions connected to the DLD outlets. Droplets act as pressure controllers, which perform at the same time the encapsulation of DLD sorted particles and the balance of output pressures. The optimized pressures to apply on DLD modules and on T-junctions are determined by a general model that ensures the equilibrium of the entire platform. The proposed separation platform is completely modular and reconfigurable since the same predictive model applies to any cascaded DLD modules of the droplet-based cartridge.