Search

Background: Long-lasting insecticidal nets (LLINs) are ineffective malaria transmission prevention tools if they are unused. Discomfort due to heat is the most commonly reported reason for not using nets, but this problem is largely unaddressed. With increasing rural electrification and the dropping price of solar power, fans could improve comfort inside nets and be affordable to populations in malaria endemic areas. Here, results are presented from a pilot randomized controlled cross-over study testing the effect of fans on LLIN use., Methods: Eighty-three households from two rural communities in Greater Accra, Ghana, randomized into three groups, participated in a 10-month cross-over trial. After a screening survey to identify eligible households, all households received new LLINs. Bͻkͻͻ net fan systems (one fan per member) were given to households in Group 1 and water filters were given to households in Group 2. At mid-point, Group 1 and 2 crossed over interventions. Households in Group 1 and 2 participated in fortnightly surveys on households' practices related to nets, fans and water filters, while households in Group 3 were surveyed only at screening, mid-point and study end. Entomological and weather data were collected throughout the study. Analysis took both 'per protocol' (PP) and 'intention to treat' (ITT) approaches. The mid- and end-point survey data from Group 1 and 2 were analysed using Firth logistic regressions. Fortnightly survey data from all groups were analysed using logistic regressions with random effects., Results: Provision of fans to households appeared to increase net use in this study. Although the increase in net use explained by fans was not significant in the primary analyses (ITT odds ratio 3.24, p > 0.01; PP odds ratio = 1.17, p > 0.01), it was significant in secondary PP analysis (odds ratio = 1.95, p < 0.01). Net use was high at screening and even higher after provision of new LLINs and with follow up. Fan use was 90-100% depending on the fortnightly visit., Conclusions: This pilot study could not provide definitive evidence that fans increase net use. A larger study with additional statistical power is needed to assess this association across communities with diverse environmental and socio-demographic characteristics.

Background: Long-lasting insecticidal nets (LLINs) are known to be highly effective in reducing malaria transmission, morbidity and mortality. However, among those owning an LLIN, use rates are often suboptimal. A reported barrier to bed net use is discomfort due to heat. This qualitative study was part of a larger evaluation conducted in communities without electricity in rural Ghana to assess whether 0.8 W solar powered net fans can increase net use., Methods: Twenty-three key informant interviews with household heads in the study communities in Shai-Osudoku District, southern Ghana, were conducted from July to August 2015. The purpose of the interviews was to obtain insight into perceptions of participants about the net fan system in relation to LLIN use., Results: While all study participants reported using LLINs, with mosquito nuisance prevention as the prime motivation, heat was also mentioned as a key barrier to net use. Respondents appreciated the net fans because they improved comfort inside bed nets. The LED light on the fan stand became the main source of light at night and positively influenced the perception of the intervention as a whole., Conclusion: The general acceptance of the net fan system by the study participants highlights the potential of the intervention to improve comfort inside mosquito nets. This, therefore, has a potential to increase bed net use in areas with low access to electricity.

Prediction of drug-induced toxicity is complicated by the failure of animal models to extrapolate human response, especially during assessment of repeated dose toxicity for cosmetic or chronic drug treatments. In this work, we present a 3D microreactor capable of maintaining metabolically active HepG2/C3A spheroids for over 28 days in vitro under stable oxygen gradients mimicking the in vivo microenvironment. Mitochondrial respiration was monitored using two-frequency phase modulation of phosphorescent microprobes embedded in the tissue. Phase modulation is focus independent and unaffected by cell death or migration. This sensitive measurement of oxygen dynamics revealed important information on the drug mechanism of action and transient subthreshold effects. Specifically, exposure to antiarrhythmic agent, amiodarone, showed that both respiration and the time to onset of mitochondrial damage were dose dependent showing a TC50 of 425 μm. Analysis showed significant induction of both phospholipidosis and microvesicular steatosis during long-term exposure. Importantly, exposure to widely used analgesic, acetaminophen, caused an immediate, reversible, dose-dependent loss of oxygen uptake followed by a slow, irreversible, dose-independent death, with a TC50 of 12.3 mM. Transient loss of mitochondrial respiration was also detected below the threshold of acetaminophen toxicity. The phenomenon was repeated in HeLa cells that lack CYP2E1 and 3A4, and was blocked by preincubation with ascorbate and TMPD. These results mark the importance of tracing toxicity effects over time, suggesting a NAPQI-independent targeting of mitochondrial complex III might be responsible for acetaminophen toxicity in extrahepatic tissues.

In this study, the effect of temperature on the build-up of exponentially growing polyelectrolyte multilayer films was investigated. It aims at understanding the multilayer growth mechanism as crucially important for the fabrication of tailor-made multilayer films. Model poly(L-lysine)/hyaluronic acid (PLL/HA) multilayers were assembled in the temperature range of 25-85 °C by layer-by-layer deposition using a dipping method. The film growth switches from the exponential to the linear regime at the transition point as a result of limited polymer diffusion into the film. With the increase of the build-up temperature the film growth rate is enhanced in both regimes; the position of the transition point shifts to a higher number of deposition steps confirming the diffusion-mediated growth mechanism. Not only the faster polymer diffusion into the film but also more porous/permeable film structure are responsible for faster film growth at higher preparation temperature. The latter mechanism is assumed from analysis of the film growth rate upon switching of the preparation temperature during the film growth. Interestingly, the as-prepared films are equilibrated and remain intact (no swelling or shrinking) during temperature variation in the range of 25-45 °C. The average activation energy for complexation between PLL and HA in the multilayers calculated from the Arrhenius plot has been found to be about 0.3 kJ mol(-1) for monomers of PLL. Finally, the following processes known to be dependent on temperature are discussed with respect to the multilayer growth: (i) polymer diffusion, (ii) polymer conformational changes, and (iii) inter-polymer interactions.

Microfluidic applications range from combinatorial synthesis to high throughput screening, with platforms integrating analog perfusion components, digitally controlled micro-valves and a range of sensors that demand a variety of communication protocols. Currently, discrete control units are used to regulate and monitor each component, resulting in scattered control interfaces that limit data integration and synchronization. Here, we present a microprocessor-based control unit, utilizing the MS Gadgeteer open framework that integrates all aspects of microfluidics through a high-current electronic circuit that supports and synchronizes digital and analog signals for perfusion components, pressure elements, and arbitrary sensor communication protocols using a plug-and-play interface. The control unit supports an integrated touch screen and TCP/IP interface that provides local and remote control of flow and data acquisition. To establish the ability of our control unit to integrate and synchronize complex microfluidic circuits we developed an equi-pressure combinatorial mixer. We demonstrate the generation of complex perfusion sequences, allowing the automated sampling, washing, and calibrating of an electrochemical lactate sensor continuously monitoring hepatocyte viability following exposure to the pesticide rotenone. Importantly, integration of an optical sensor allowed us to implement automated optimization protocols that require different computational challenges including: prioritized data structures in a genetic algorithm, distributed computational efforts in multiple-hill climbing searches and real-time realization of probabilistic models in simulated annealing. Our system offers a comprehensive solution for establishing optimization protocols and perfusion sequences in complex microfluidic circuits.

Understanding the dynamics of signal transduction processes that are induced by cell-cell or cell-surface interactions requires the physical stimulation of the cells of interest on a single-cell level and without any ill-defined contacting of their cell membrane. However, standard cell culture techniques are inapplicable for this task as they do not provide cell and particle handling at sufficiently high spatial and temporal resolution and are limited to ensemble measurements. Here, we present a novel process line for the individual stimulation of single cells with bioactive surfaces, like other cells or particles, and the simultaneous analysis of the induced cytosolic calcium signals. The method is based on a microfluidic lab-on-a-chip environment that allows for contactless cell and particle handling by dielectrophoretic forces.

Monitoring cellular bioenergetic pathways provides the basis for a detailed understanding of the physiological state of a cell culture. Therefore, it is widely used as a tool amongst others in the field of in vitro toxicology. The resulting metabolic information allows for performing in vitro toxicology assays for assessing drug-induced toxicity. In this study, we demonstrate the value of a microsystem for the fully automated detection of drug-induced changes in cellular viability by continuous monitoring of the metabolic activity over several days. To this end, glucose consumption and lactate secretion of a hepatic tumor cell line were continuously measured using microfluidically addressed electrochemical sensors. Adapting enzyme-based electrochemical flat-plate sensors, originally designed for human whole-blood samples, to their use with cell culture medium supersedes the common manual and laborious colorimetric assays and off-line operated external measurement systems. The cells were exposed to different concentrations of the mitochondrial inhibitor rotenone and the cellular response was analyzed by detecting changes in the rates of the glucose and lactate metabolism. Thus, the system provides real-time information on drug-induced liver injury in vitro.

The prospect of novel therapeutic approaches has renewed the current interest in the fusion of rare cells, like stem cells or primary immune cells. While conventional techniques are only capable of mass fusion, lab-on-a-chip systems often still lack an acceptable method for making the cells available after processing. Here, we present a microfluidic approach for electrofusion on the single-cell level that offers high control over the cells both before and after fusion. For cell pairing and fusion, we employed dielectrophoresis and AC voltage pulses, respectively. Each cell has been characterized and selected before they were paired, fused and released from the fluidic system for subsequent analysis and cultivation. The successful experimental evaluation of our system was further corroborated by numerical simulations. We obtained fusion efficiencies of more than 30% for individual pairs of mouse myeloma and B cell blasts and showed the proliferating ability of the hybrid cells 3 d after fusion. Since aggregates of more than two cells can be fused, the technique could also be developed further for generating giant cells for low-noise electrophysiology in the context of semi-automated pharmaceutical screening procedures.

We present an electrohydrodynamic device for filtration of nanometre-sized particles from suspensions. A high-frequency electric field is locally generated through the action of mutually parallel microelectrodes integrated into a microfluidic channel. Due to the mechanism of ohmic heating, a thermal gradient arises above these electrodes. In conjunction with temperature-sensitive properties of the fluid, an eddy flow behaviour emerges in the laminar environment. This acts as an adjustable filter. For quantification of the filtration efficiency, we tested a wide range of particle concentrations at different electric field strengths and overall external flow velocities. Particles with a diameter of 200 nm were retained in this manner at rates of up to 100%. Numerical simulations of a model taking into account the hydrodynamic as well as electric conditions, but no interactions between the point-shaped particles, yield results that are similar to the experiment in both the flow trajectories and the particle accumulation. Our easy technique could become a valuable tool that complements conventional filtration methods for handling nanometre-scaled particles in medicine and biotechnology, e.g. bacteria and viruses.

The mechanical behavior of biological cells is mainly determined by the cytoskeleton. Its properties are closely interlinked with many cellular events, including disease-related processes, and, thus, may be exploited as potent biomarkers. We have stretched two types of cells between microelectrodes through the application of dielectrophoretic forces. Small numbers of cells of cancerous origin (MCF-7) and from related noncancerous tissue (MCF-10A) were sufficient to obtain data that allowed us to unambiguously distinguish these cells. The Maxwell tension applied has been estimated to be 56 Pa. A detailed analysis of the cells showed that the differences in the stretching response are due to cell-specific mechanical properties. Through the addition of an actin- and a microtubule-specific toxin to the cells, differences in the microtubular structures of the two cell types have been identified as the major cause for the behavior observed. Our approach shows enormous potential for parallelization and automation. Hence, it should be suitable for achieving throughputs that make it attractive for many biomedical diagnostic purposes.

The mechanical behavior of a living cell is highly dynamic and constantly adapts to its local environment. Changes in temperature and chemical stimuli, such as pH, may alter the structure of the cell and its mechanical response. Thus, the mechanical properties may serve as an indicator for the cellular state. We applied dielectrophoretic forces to suspension cells by means of two microelectrodes. The resultant stretching was analyzed on consecutive cultivation days with respect to the influence of medium consumption. Systematic experiments clearly showed that the medium consumption affected the viscoelastic properties of the investigated human leukemia cells HL-60. The shift in pH value and the culture medium depletion were identified as potentially responsible for the differing temporal development of the cell deformation. Both factors were investigated separately and a detailed analysis indicated that the changes observed in the cellular stiffness were primarily attributable to nutrient depletion.

In order to elucidate the dynamics of cellular processes that are induced in context with intercellular communication, defined events along the signal transduction cascade and subsequent activation steps have to be analyzed on the level of individual cells and correlated with each other. Here we present an approach that allows the initiation of cell-cell or cell-particle interactions and the analysis of cellular reactions within various regimes while the identity of each individual cell is preserved. It utilizes dielectrophoresis (DEP) and microfluidics in a lab-on-chip system. With high spatial and temporal precision we contacted single T cells with functionalized microbeads and monitored their immediate cytosolic Ca(2+) response. After this, the cells were released from the chip system and cultivated further. Expression of the activation marker molecule CD69 was analyzed the next day and correlated with the previously recorded Ca(2+) signal for each individual cell. We found a significant difference in the patterns of Ca(2+) traces between activated and non-activated cells, which shows that Ca(2+) signals in T cells can provide early information about a later reaction of the cell. Although T cells are non-excitable cells, we also observed irregular Ca(2+) transients upon exposure to the DEP field only. These Ca(2+) signals depended on exposure time, electric field strength and field frequency. By minimizing their occurrence rate, we could identify experimental conditions that caused the least interference with the physiology of the cell.

The gentle and careful in vitro processing of live cells is essential in order to make them available to future therapeutic applications. We present a protocol for the activation of single-T cells based on the contact formation with individual anti-CD3/anti-CD28 presenting microbeads in a lab-on-chip environment. The chips consist of microfluidic channels and microelectrodes for performing dielectrophoretic manipulation employing a.c. electric fields. The dielectrophoretic guiding elements allow the assembly of cell-bead pairs while avoiding ill-defined physical contacts with their environment. After overnight cultivation of the manipulated cells, 77% of the bead-associated T cells expressed the activation marker molecule CD69. Physiological stress on the cells was shown to be mainly due to the single-cell cultivation and not to the manipulation in the chips. The same approach could be useful for the in vitro regulation of stem cell differentiation.

We present an approach for the accumulation and filtering of nano- and microparticles in microfluidic devices that is based on the generation of electric traveling waves in the radio-frequency range. Upon application of the electric field via a microelectrode array, complex particle trajectories and particle accumulation are observed in well-defined regions in a microchannel. Through the quantitative mapping of the 3-D flow pattern using two-focus fluorescence cross-correlation spectroscopy, two vortices could be identified as one of the sources of the force field that induces the formation of particle clouds. Dielectrophoretic forces that directly act on the particles are the second source of the force field. A thorough 2-D finite element analysis identifies the electric traveling wave mechanism as the cause for the unexpected flow behavior observed. Based on these findings, strategies are discussed, first, for avoiding the vortices to optimize electrohydrodynamic micropumps and, secondly, for utilizing the vortices in the development of microdevices for efficient particle accumulation, separation, and filtering. Such devices may find numerous biomedical applications when highly diluted nano- and microsuspensions have to be processed.

Although they have been discovered decades ago, only in the last years forisome protein aggregates received broader attention due to their ability to convert chemical into mechanical energy. In contrast to most other motor proteins, these proteins from Fabaceae plants are independent of high-energy chemical compounds, like e.g. ATP, but undergo an anisotropic shape transition (longitudinally expanded to contracted) in response to ion concentration changes (Ca(2+), H(+), etc.), instead. We present morphological and functional data on forisomes obtained using atomic force microscopy (AFM). High-aspect ratio AFM tips allow the detailed elucidation of structural characteristics that are inaccessible with standard AFM tips. Microindentation measurements were employed to calculate the elasticity of the forisome material. Young's moduli were found to be approximately 32.7 kPa in the expanded state and approximately 2.748 kPa in the contracted state of the polymer. These results are compared to investigations where a tipless AFM cantilever was utilized to exert a load against the shape transition. In the latter experiments, an energy conversion of approximately 2.29 pJ per stroke was detected. Energetical considerations support the hypothesis that the switching process is accompanied by a change in cross-linking of the constituent subunits and allow estimating the extent of cooperativity during the pH-induced transition. Finally, useful parameters were identified and characterized that are crucial for the application of forisomes as functional elements in microfluidic chips.