Single Cell Bioprinting with Ultrashort Laser Pulses

Tissue engineering requires the precise positioning of mammalian cells and biomaterials on substrate surfaces or in preprocessed scaffolds. Although the development of 2D and 3D bioprinting technologies has made substantial progress in recent years, precise, cell-friendly, easy to use, and fast technologies for selecting and positioning mammalian cells with single cell precision are still in need. A new laser-based bioprinting approach is therefore presented, which allows the selection of individual cells from complex cell mixtures based on morphology or fluorescence and their transfer onto a 2D target substrate or a preprocessed 3D scaffold with single cell precision and high cell viability (93–99% cell survival, depending on cell type and substrate). In addition to precise cell positioning, this approach can also be used for the generation of 3D structures by transferring and depositing multiple hydrogel droplets. By further automating and combining this approach with other 3D printing technologies, such as two-photon stereolithography, it has a high potential of becoming a fast and versatile technology for the 2D and 3D bioprinting of mammalian cells with single cell resolution.     

Alloy Design,Combinatorial Synthesis,and Microstructure–Property Relations for Low-Density Fe-Mn-Al-C Austenitic Steels

We present recent developments in the field of austenitic steels with up to 18% reduced mass density. The alloys are based on the Fe-Mn-Al-C system. Here, two steel types are addressed. The first one is a class of low-density twinning-induced plasticity or single phase austenitic TWIP (SIMPLEX) steels with 25–30 wt.% Mn and <4–5 wt.% Al or even <8 wt.% Al when naturally aged. The second one is a class of κ-carbide strengthened austenitic steels with even higher Al content. Here, κ-carbides form either at 500–600°C or even during quenching for >10 wt.% Al. Three topics are addressed in more detail, namely, the combinatorial bulk high-throughput design of a wide range of corresponding alloy variants, the development of microstructure–property relations for such steels, and their susceptibility to hydrogen embrittlement.     

Numerical and experimental methodology for the development of a new membrane prototype intended to microfiltration bioprocesses. Application to milk filtration

In tangential flow filtration, the non-uniform TransMembrane Pressure (TMP) on the membrane length produces a non homogeneous filtration cake, initiates process selectivity changes and modifies the permeate quality. The purpose of this study is to create a tubular ceramic membrane prototype with a more uniform TMP, intended to filtration of fouling fluids. The principle of this membrane structure is to waterproof the external membrane surface to limit flow circulation in the porous support of the membrane. The production was controlled by sizing «permeation vents». This development was achieved using a CFD modelling tool interacting with experiments. A preliminary modelling study was made with water. This work was afterwards applied to the industrial process of casein micelle separation from skim milk. The influence of operating conditions on the membrane hydrodynamics was highlighted. The modelling results were experimentally confirmed, with a discrepancy smaller than 3% and a reproducible water permeability of 2.3 L h⁻¹ bar⁻¹ for 1 mm-wide vent (TMP = 1 bar, T = 20 °C). Then, milk filtration experiments showed a production ratio milk/water equal to 1/2. The permeate quality parameters were studied and the fouling phenomena were taken into account. A parametric study led to the sizing of a final prototype. Its efficiency was experimentally evaluated.     

Mechanical and electric fatigue of PZT(53/47) films on metallic substrates

Abstract

We have investigated the fatigue of electromechanical and dielectric properties of sol-gel derived PZT(53/47) thin films deposited on metallic substrates by means of electric and mechanical cycling. For the mechanical cycling a two point bending method was used to apply transversal stress to the samples. During mechanical cycling the piezoelectric coefficient d₃₁ remained constant up to about 10⁵ cycles, for a higher number of cycles a strong decrease was observed. During electric cycling no significant changes in the ferroelectric and electromechanical hysteresis loops could be found up to about 3×10⁵ cycles. Above this number the coercive field increases, the maximum strain and the remanent polarization decrease.

Obviously each electric cycling of the investigated films is accompanied by a mechanical cycling. It is assumed, that microcracks induced by mechanical stress are the main reason for the deterioration of the physical properties films during electric and mechanical cycling both.     

A Fully Automated Process Using a Magnetic Particle Based Kit for Removal of Dye Terminators from Sequencing Reactions

Prolinx,® Inc. of Bothell, WA has developed the RapXtract™ 384 Dye Terminator Removal Kit for full automation of DNA sequencing reaction purification. The RapXtract product line is based upon proprietary superparamagnetic particle technology that eliminates the need for centrifugation, vacuum filtration, or modified primers to achieve purification of sequencing reactions. The kit described here is pre-dispensed in a 384-well microtiter plate and run on the TECAN GENESIS Workstation 150 (Tecan U.S. Inc., Research Triangle Park, NC). This system enables rapid purification of up to 384 sequencing reactions in a single run.As the completion of the Human Genome Project nears, it is imperative for biotechnology and pharmaceutical companies to increase throughput of DNA sequencing in order to be competitive in the drug discovery and validation process. The “race to market” requires a shift from standard DNA sequencing processes-including DNA sequencing reaction purification-towards complete walk-away automation for all steps.Existing sequencing reaction purification methods (Table 1) require considerable resources including: plastic and other laboratory consumables; specialized equipment, such as high-speed centrifuges or vacuum filtration apparatus; and labor-intensive protocols requiring large amounts of technician time. As a result, walk-away automation of standard purification methods is difficult and expensive.     

An automatic braking system that stabilizes leukocyte rolling by an increase in selectin bond number with shear

Wall shear stress in postcapillary venules varies widely within and between tissues and in response to inflammation and exercise. However, the speed at which leukocytes roll in vivo has been shown to be almost constant within a wide range of wall shear stress, i.e., force on the cell. Similarly, rolling velocities on purified selectins and their ligands in vitro tend to plateau. This may be important to enable rolling leukocytes to be exposed uniformly to activating stimuli on endothelium, independent of local hemodynamic conditions. Wall shear stress increases the rate of dissociation of individual selectin-ligand tether bonds exponentially (, ) thereby destabilizing rolling. We find that this is compensated by a shear-dependent increase in the number of bonds per rolling step. We also find an increase in the number of microvillous tethers to the substrate. This explains (a) the lack of firm adhesion through selectins at low shear stress or high ligand density, and (b) the stability of rolling on selectins to wide variation in wall shear stress and ligand density, in contrast to rolling on antibodies (). Furthermore, our data successfully predict the threshold wall shear stress below which rolling does not occur. This is a special case of the more general regulation by shear of the number of bonds, in which the number of bonds falls below one.