On chip electrofusion of single human B cells and mouse myeloma cells for efficient hybridoma generation

This article describes the development and full characterization of a microfluidic chip for electrofusion of human peripheral blood B-cells and mouse myeloma (NS-1) cells to generate hybridomas. The chip consists of an array of 783 traps, with dimensions that were optimized to obtain a final cell pairing efficiency of 33±6%. B cells were stained with a cytoplasmic stain CFDA to assess the different stages of cell fusion, i.e. dye transfer to NS-1 cells (initiating fusion) and membrane reorganization (advanced fusion). Six DC pulses of 100 μs (2.5 kV/cm) combined with an AC field (30 s, 2 MHz, 500 V/cm) and pronase treatment resulted in the highest electrofusion efficiency of paired cells (51±11%). Hybridoma formation, with a yield of 0.33 and 1.2%, was observed after culturing the fused cells for 14 days in conditioned medium. This work provides valuable leads to improve the current electrofusion protocols for the production of human antibodies for diagnostic and therapeutic applications.     

Field and pressure response of Yb compounds close to a quantum critical point

YbCu_5−x Al _x provides the possibility to tune ground state properties by a change of the valence due to the Cu/Al substitution, by pressure as well as by the application of a magnetic field. Near to the critical concentration x _cr≈1.5 non-Fermi-liquid properties (NFL) are obvious, obeying hyperscaling. If magnetic order sets in for x>1.5, the application of moderate magnetic fields quenches order and again NFL features become evident. Hyperscaling in this case indicates strongly interacting spin fluctuations.     

Complex conductivity of water-saturated packs of glass beads

The low-frequency conductivity response of water-saturated packs of glass beads reflects a combination of two processes. One process corresponds to the polarization of the mineral/water interface coating the surface of the grains. The other process corresponds to the Maxwell-Wagner polarization associated with accumulation of the electrical charges in the pore space of the composite medium. A model of low-frequency conductivity dispersion is proposed. This model is connected to a triple-layer model of electrochemical processes occurring at the surface of silica. This model accounts for the partition of the counterions between the Stern and the diffuse layers. The polarization of the mineral/water interface is modeled by the electrochemical polarization model of Schurr for a spherical grain. We take into account also the DC surface conductivity contribution of protons of the sorbed water and the contribution of the diffuse layer. At the scale of a macroscopic representative elementary volume of the porous material, the electrochemical polarization of a single grain is convoluted with the grain size distribution of the porous material. Finally, the Maxwell-Wagner polarization is modeled using the complex conductivity of a granular porous medium obtained from the differential effective medium theory. The predictions of this model agree well with experimental data of spectral induced polarization. Two peaks are observed at low frequencies in the spectrum of the phase. The first peak corresponds to the distribution of the size of the beads and the second peak is due to the roughness of the grains.     

Enzymatic Crosslinking of Polymer Conjugates is Superior over Ionic or UV Crosslinking for the On‐Chip Production of Cell‐Laden Microgels

Cell‐laden micrometer‐sized hydrogels (microgels) hold great promise for improving high throughput ex‐vivo drug screening and engineering biomimetic tissues. Microfluidics is a powerful tool to produce microgels. However, only a limited amount of biomaterials have been reported to be compatible with on‐chip microgel formation. Moreover, these biomaterials are often associated with mechanical instability, cytotoxicity, and cellular senescence. To resolve this challenge, dextran‐tyramine has been explored as a novel biomaterial for on‐chip microgel formation. In particular, dextran‐tyramine is compared with two commonly used biomaterials, namely, polyethylene‐glycol diacrylate (PEGDA) and alginate, which crosslink through enzymatic reaction, UV polymerization, and ionic interaction, respectively. Human mesenchymal stem cells (hMSCs) encapsulated in dextran‐tyramine microgels demonstrate significantly higher (95%) survival as compared to alginate (81%) and PEGDA (69%). Long‐term cell cultures demonstrate that hMSCs in PEGDA microgels become senescent after 7 d. Alginate microgels dissolve within 7 d due to Ca²⁺ loss. In contrast, dextran‐tyramine based microgels remain stable, sustain hMSCs metabolic activity, and permit for single‐cell level analysis for at least 28 d of culture. In conclusion, enzymatically crosslinking dextran‐tyramine conjugates represent a novel biomaterial class for the on‐chip production of cell‐laden microgels, which possesses unique advantages as compared to the commonly used UV and ionic crosslinking biomaterials.

     

High-yield cell ordering and deterministic cell-in-droplet encapsulation using Dean flow in a curved microchannel

In this article high-yield (77%) and high-speed (2700 cells s(-1)) single cell droplet encapsulation is described using a Dean-coupled inertial ordering of cells in a simple curved continuous microchannel. By introducing the Dean force, the particles will order to one equilibrium position after travelling less than 1 cm. We use a planar curved microchannel structure in PDMS to spatially order two types of myeloid leukemic cells (HL60 and K562 cells), enabling deterministic single cell encapsulation in picolitre drops. An efficiency of up to 77% was reached, overcoming the limitations imposed by Poisson statistics for random cell loading, which yields only 37% of drops containing a single cell. Furthermore, we confirm that > 90% of the cells remain viable. The simple planar structure and high throughput provided by this passive microfluidic approach makes it attractive for implementation in lab on a chip (LOC) devices for single cell applications using droplet-based platforms.