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By considering diffusion, convection (the flow along a channel) and capillary forces, the authors designed a microfluidic structure that produces a complicated, yet highly reproducible, liquid flow that first passes around dried spots of reagents and then back over them. The ratio of the surface area of a microchannel- confined liquid (that is, the surface area bounded by the channel walls) to its volume is large, allowing heat and mass to be rapidly transferred to such liquids. [...]the flow of the liquid can be tightly controlled. The authors went on to demonstrate how their system could be used to precisely control the concentration and the timing of addition of reagents in complex biochemical reactions, in two assays: one that detected DNA sequences of the human papilloma virus, and the other that quantified the activity of an enzyme.

Microfluidic devices have revolutionized biological assays, but complex set-ups are required to prevent the unwanted mixing of reagents in the liquid samples being analysed. A simpler solution has just been found. How to prevent unwanted dispersal of reagents in microfluidic systems.

Nanoviscosity of the cytoplasm is a key factor affecting diffusion of biomolecules and – as a consequence – rates of biochemical reactions in a cell. Nanoviscosity is an outcome of variable chemical and structural factors, which can temporarily change with cell-cycle associated changes of intracellular architecture. Thus, the question arises, whether rates of biochemical reactions depend on the point of cell cycle. In this paper we address this topic by constant observation of nanoviscosity of HeLa cells cytoplasm during S, G2 and G1 phases after Aphidicolin synchronization. For this purpose we measured diffusion rates of EGFP molecules using fluorescence correlation spectroscopy (FCS). To our surprise, a counter-intuitive stability of cytoplasmic viscosity was observed during the cell cycle. Our results hint at possible existence of robust mechanism maintaining stable physiological viscosity of the cytoplasm, despite huge structural changes during cell cycle.

Hydrophobicity is one of the most critical factors governing the adsorption of molecules and objects, such as virions, on surfaces. Even moderate change of wetting angle of plastic surfaces causes a drastic decrease ranging from 2 to 5 logs of the viruses (e.g., T4 phage) in the suspension due to adsorption on polymer vials' walls. The effect varies immensely in seemingly identical containers but purchased from different vendors. Comparison of glass, polyethylene, polypropylene, and polystyrene containers revealed a threshold in the wetting angle of around 95°: virions adsorb on the surface of more hydrophobic containers, while in more hydrophilic vials, phage suspensions are stable. The polypropylene surface of the Eppendorf-type and Falcon-type can accommodate from around 10 8 PFU/ml to around 10 10 PFU/ml from the suspension. The adsorption onto the container’s wall might result in complete scavenging of virions from the bulk. We developed two methods to overcome this issue. The addition of surfactant Tween20 and/or plasma treatment provides a remedy by modulating surface wettability and inhibiting virions' adsorption. Plastic containers are essential consumables in the daily use of many bio-laboratories. Thus, this is important not only for phage-related research (e.g., the use of phage therapies as an alternative for antibiotics) but also for data comparison and reproducibility in the field of biochemistry and virology.

Trimethylamine-N-oxide (TMAO) has been suggested as a marker and mediator of cardiovascular diseases. However, data are contradictory, and the mechanisms are obscure. Strikingly, the role of the TMAO precursor trimethylamine (TMA) has not drawn attention in cardiovascular studies even though toxic effects of TMA were proposed several decades ago. We assessed plasma TMA and TMAO levels in healthy humans (HH) and cardiovascular patients qualified for aortic valve replacement (CP). The cytotoxicity of TMA and TMAO in rat cardiomyocytes was evaluated using an MTT test. The effects of TMA and TMAO on albumin and lactate dehydrogenase (LDH) were assessed using fluorescence correlation spectroscopy. In comparison to HH, CP had a two-fold higher plasma TMA (p < 0.001) and a trend towards higher plasma TMAO (p = 0.07). In CP plasma, TMA was inversely correlated with an estimated glomerular filtration rate (eGFR, p = 0.002). TMA but not TMAO reduced cardiomyocytes viability. Incubation with TMA but not TMAO resulted in the degradation of the protein structure of LDH and albumin. In conclusion, CP show increased plasma TMA, which is inversely correlated with eGFR. TMA but not TMAO exerts negative effects on cardiomyocytes, likely due to its disturbing effect on proteins. Therefore, TMA but not TMAO may be a toxin and a marker of cardiovascular risk.

The oxazole yellow dye, YOYO-1 (a symmetric homodimer), is a commonly used molecule for staining DNA. We applied the brightness analysis to study the intercalation of YOYO-1 into the DNA. We distinguished two binding modes of the dye to dsDNA: mono-intercalation and bis-intercalation. Bis-intercalation consists of two consecutive mono-intercalation steps, characterised by two distinct equilibrium constants (with the average number of base pair per binding site equals 3.5): K1=3.36±0.43×107M−1 and K2=1.90±0.61×105M−1, respectively. Mono-intercalation dominates at high concentrations of YOYO-1. Bis-intercalation occurs at low concentrations.

There is a long-standing question of whether it is possible to extend the formalism of equilibrium thermodynamics to the case of nonequilibrium systems in steady-states. We have made such an extension for an ideal gas in a heat flow. Here, we investigated whether such a description exists for the system with interactions: the van der Waals gas in a heat flow. We introduced a steady-state fundamental relation and the parameters of state, each associated with a single way of changing energy. The first law of nonequilibrium thermodynamics follows from these parameters. The internal energy U for the nonequilibrium states has the same form as in equilibrium thermodynamics. For the van der Waals gas, U(S*,V,N,a*,b*) is a function of only five parameters of state (irrespective of the number of parameters characterizing the boundary conditions): the effective entropy S*, volume V, number of particles N, and rescaled van der Waals parameters a*, b*. The state parameters, a*, b*, together with S*, determine the net heat exchange with the environment. The net heat differential does not have an integrating factor. As in equilibrium thermodynamics, the steady-state fundamental equation also leads to the thermodynamic Maxwell relations for measurable steady-state properties.

The reaction kinetics between like-charged compounds in water is extremely slow due to Coulomb repulsions. Here, we demonstrate that by screening these interactions and, in consequence, increasing the local concentration of reactants, we boost the reactions by many orders of magnitude. The reaction between negatively charged Coenzyme A molecules accelerates ~5 million-fold using cationic micelles. That is ~10 4 faster kinetics than in 0.5 M NaCl, although the salt is ~10 6 more concentrated. Rate enhancements are not limited to micelles, as evidenced by significant catalytic effects (10 4 –10 5 -fold) of other highly charged species such as oligomers and polymers. We generalize the observed phenomenon by analogously speeding up a non-covalent complex formation—DNA hybridization. A theoretical analysis shows that the acceleration is correlated to the catalysts’ surface charge density in both experimental systems and enables predicting and controlling reaction rates of like-charged compounds with counter-charged species. The reaction speed of like-charged compounds in water is extremely slow due to Coulomb repulsions. Here, the authors boost kinetics up to 5 million times by screening these interactions and increasing the local concentration of reactants using positively charged micelles. They show the effect for two independent systems and present a theoretical explanation.

We showed that the Taylor Dispersion Analysis (TDA) is a fast and easy to use method for the study of denaturation proteins. We applied TDA to study denaturation of β-lactoglobulin, transferrin, and human insulin by anionic surfactant sodium dodecyl sulfate (SDS). A series of measurements at constant protein concentration (for transferrin was 1.9 x 10-5 M, for β- lactoglobulin was 7.6 x 10-5 M, and for insulin was 1.2 x 10-4 M) and varying SDS concentrations were carried out in the phosphate-buffered saline (PBS). The structural changes were analyzed based on the diffusion coefficients of the complexes formed at various surfactant concentrations. The concentration of surfactant was varied in the range from 1.2 x 10-4 M to 8.7 x 10-2 M. We determined the minimum concentration of the surfactant necessary to change the native conformation of the proteins. The minimal concentration of SDS for β-lactoglobulin and transferrin was 4.3 x 10-4 M and for insulin 2.3 x 10-4 M. To evaluate the TDA as a novel method for studying denaturation of proteins we also applied other methods i.e. electronic circular dichroism (ECD) and dynamic light scattering (DLS) to study the same phenomenon. The results obtained using these methods were in agreement with the results from TDA.

Trimethylamine-oxide (TMAO) is present in seafood which is considered to be beneficial for health. Deep-water animals accumulate TMAO to protect proteins, such as lactate dehydrogenase (LDH), against hydrostatic pressure stress (HPS). We hypothesized that TMAO exerts beneficial effects on the circulatory system and protects cardiac LDH exposed to HPS produced by the contracting heart. Male, Sprague-Dawley and Spontaneously-Hypertensive-Heart-Failure (SHHF) rats were treated orally with either water (control) or TMAO. In vitro, LDH with or without TMAO was exposed to HPS and was evaluated using fluorescence correlation spectroscopy. TMAO-treated rats showed higher diuresis and natriuresis, lower arterial pressure and plasma NT-proBNP. Survival in SHHF-control was 66% vs 100% in SHHF-TMAO. In vitro, exposure of LDH to HPS with or without TMAO did not affect protein structure. In conclusion, TMAO reduced mortality in SHHF, which was associated with diuretic, natriuretic and hypotensive effects. HPS and TMAO did not affect LDH protein structure. Heart failure is a common cause of death in industrialized countries with aging populations. Japan, however, has lower rates of heart failure and fewer deaths linked to this disease than the United States or Europe, despite having the highest proportion of elderly people in the world. Dietary differences between these regions may explain the lower rate of heart failure in Japan. The Japanese diet is rich in seafood, which contains nutrients that promote heart health, such as omega-3 fatty acids. Seafood also contains other compounds, including trimethylamine oxide (TMAO). Fish that live in deep waters undergo high pressures, which can damage their proteins, but TMAO seems to protect the proteins from harm. In humans, eating seafood increases TMAO levels in the blood and urine, but it is unclear what effects this has on heart health. Increased levels of TMAO in the blood are associated with cardiovascular diseases, but scientists are not sure whether TMAO itself harms the heart. A toxic byproduct of gut bacteria called TMA is converted in TMAO in the body, so it is possible that TMA rather than TMAO is to blame. To assess the effects of dietary TMAO on heart failure, Gawrys-Kopczynska et al. fed the compound to healthy rats and rats with heart failure for one year. TMAO had no effects on the healthy rats. Of the rats with heart failure that were fed TMAO, all of them survived the year, while one third of rats with heart failure that were not fed TMAO died. TMAO-treated rats with heart failure had lower blood pressure and urinated more than untreated rats with the condition. The experiments suggest that dietary TMAO may mimic the effects of heart failure treatments, which remove excess water and salt and lower pressure on the heart. More studies are needed to confirm whether TMAO has this same effect on humans.