High-throughput screening-compatible single-step protocol to differentiate embryonic stem cells in neurons.

Biotechnologies such as high-throughput screening (HTS) enable evaluation of large compound libraries for their biological activity and toxic properties. In the field of drug development, embryonic stem (ES) cells have been instrumental in HTS for testing the effect of new compounds. We report an innovative method in one step to differentiate ES cells in neurons and glial cells. The four different neuronal subtypes, gamma-aminobutyric acid (GABA)-ergic, dopaminergic, serotonergic, and motor neurons, are formed in culture. This protocol is adaptable to small wells and is highly reproducible, as indicated by the Z-factor value. Moreover, by using either leukemia inhibitory factor (LIF) or recombinant Cripto protein in our culture conditions, we provide evidence that this protocol is suitable for testing the effect of different molecules on neuronal differentiation of ES cells. Finally, thanks to the simplicity in carrying out the experiment, this method provides the possibility of following the morphological evolution of the in vitro differentiating neuronal cells by timelapse videomicroscopy. Our experimental system provides a powerful tool for testing the effect of different substances on survival and/or differentiation of neuronal and glial cells in an HTS-based approach. Furthermore, using genetically modified ES cells, it would be possible to screen for drugs that have a therapeutic effect on specific neuronal pathologies or to follow, by time-lapse videomicroscopy, their ability to in vitro differentiate.

Shear banding in biphasic liquid-liquid systems.

In this Letter, we present the first systematic report of shear-induced banding in microconfined biphasic liquid-liquid systems, i.e., formation of alternating regions of high and low volume fraction of dispersed-phase droplets in a parallel plate flow cell. Such a flow-driven, gap-dependent phenomenon is only observed at low values of the viscosity ratio between the dispersed and the continuous phase, and in a given range of the applied shear rate. Based on rheological measurements, band formation is found to be associated with a viscosity decrease as compared to the homogeneous, structureless case, thus showing that system microstructure is somehow evolving towards reduced viscous dissipation under shear flow.

A methodology to study the deformability of red blood cells flowing in microcapillaries in vitro.

The deformability of red blood cells flowing in microvessels is essential to maintain optimal blood circulation and to allow gas transfer between blood and tissues. Here, we report on an experimental methodology to investigate the deformability of RBCs flowing in microcapillaries having diameter close to the average cell size. The microcapillaries are placed in a rectangular flow cell, where a suspension of RBCs, properly diluted in albumin-additioned ACD, is fed through a syringe under the action of a liquid head in the physiological range. Video microscopy images of the flowing RBCs are acquired at high magnification and later processed by an automated image analysis macro. It was found that RBCs from healthy donors exhibit the classical parachute shape observed in vivo. Furthermore, all the data of healthy RBC velocity vs liquid head are well represented by the same linear regression, independently on the donor. Preliminary results on beta-thalassemia RBCs are also presented and show, on the average, a reduced velocity compared to healthy samples.

Drop deformation in microconfined shear flow.

The deformation and breakup of a drop in an immiscible equiviscous liquid undergoing unbounded shear flow has been extensively investigated in the literature, starting from the pioneering work of Taylor. In this Letter, we address the case of microconfined shear flow, a problem which is relevant for microfluidics and emulsion processing applications. The main effects of confinement include complex oscillating transients and drop stabilization against breakup. In particular, very elongated drop shapes are observed, which would be unstable in the unbounded case and can be explained in terms of wall-induced distortion of the shear flow field. We show that wall effects can be exploited to obtain nearly monodisperse emulsions in microconfined shear flow.