A method for matching the refractive index and kinematic viscosity of a blood analog for flow visualization in hydraulic cardiovascular models

In this work, we propose a simple method to simultaneously match the refractive index and kinematic viscosity of a circulating blood analog in hydraulic models for optical flow measurement techniques (PIV, PMFV, LDA, and LIF). The method is based on the determination of the volumetric proportions and temperature at which two transparent miscible liquids should be mixed to reproduce the targeted fluid characteristics. The temperature dependence models are a linear relation for the refractive index and an Arrhenius relation for the dynamic viscosity of each liquid. Then the dynamic viscosity of the mixture is represented with a Grunberg-Nissan model of type 1. Experimental tests for acrylic and blood viscosity were found to be in very good agreement with the targeted values (measured refractive index of 1.486 and kinematic viscosity of 3.454 milli-m2/s with targeted values of 1.47 and 3.300 milli-m2/s).     

Microplate assay for endo-polygalacturonase activity determination based on ruthenium red method

Endo-polygalacturonase (endo-PGase) activity determinations generally rely on viscosity changes or reducing sugar ends produced by this activity over polygalacturonic acid. Torres and coworkers [Enzyme Microb. Technol. 48 (2011) 123–128] showed that ruthenium red (RR) is useful for endo-PGase determination. In this article, we present a high-throughput liquid-based endo-PGase assay based on the RR method and compare it with the viscosity determination method. The reduced assay uses a small volume of enzyme solution, 40 μg of polygalacturonic acid, and 45 μg of RR for each sample determination. Furthermore, we obtained an interconversion factor for RR and viscosity activities.     

Assessing the use of ellipsoidal microparticles for determining lipid membrane viscosity

The viscosity of lipid membranes sets the timescales of membrane-associated motions, whether driven or diffusive, and therefore influences the dynamics of a wide range of cellular processes. Techniques to measure membrane viscosity remain sparse, however, and reported measurements to date, even of similar systems, give viscosity values that span orders of magnitude. To address this, we improve a method based on measuring both the rotational and translational diffusion of membrane-anchored microparticles and apply this approach and one based on tracking the motion of phase-separated lipid domains to the same system of phase-separated giant vesicles. We find good agreement between the two methods, with inferred viscosities within a factor of two of each other. Our single-particle tracking technique uses ellipsoidal microparticles, and we show that the extraction of physically meaningful viscosity values from their motion requires consideration of their anisotropic shape. The validation of our method on phase-separated membranes makes possible its application to other systems, which we demonstrate by measuring the viscosity of bilayers composed of lipids with different chain lengths ranging from 14 to 20 carbon atoms, revealing a very weak dependence of two-dimensional viscosity on lipid size. The experimental and analysis methods described here should be generally applicable to a variety of membrane systems, both reconstituted and cellular.     

Interaction of fluorescent molecular rotors with blood plasma proteins

Many disease states have associated blood viscosity changes. Molecular rotors, fluorescent molecules with viscosity sensitive quantum yields, have recently been investigated as a new method for biofluid viscosity measurement. Current viscometer measurements are complicated by proteins adhering to surfaces and forming air-surface layers. It is unknown at this time what effects proteins may have on biofluid viscosity measurements using molecular rotors. To answer this question, binding affinities to blood plasma proteins were investigated by equilibrium dialysis for four hydrophilic molecular rotors. Aqueous solutions of 9-[(2-cyano-2-hydroxy-carbonyl)vinyl]julolidine (CCVJ) and three derivatives were prepared and dialyzed against solutions of bovine source albumin, fibrinogen and immunoglobulin G approximating normal physiologic concentrations and fresh-frozen human plasma. After equilibration, dye concentration on each side of the dialysis membrane was assessed by spectrophotometry. The relative binding affinity of the four dyes to the proteins and to the plasma was compared. Affinity of all dyes was highest for albumin. The bound dye fraction showed little change in relation to protein concentration in the physiological concentration range. Diol, the most hydrophilic molecular rotor tested showed the lowest affinity for albumin. This study indicates that hydrophilic molecular rotors are well-suited for biofluid viscosity measurement.     

Parametric dependence on shear viscosity of SPC/E water from equilibrium and non-equilibrium molecular dynamics simulations

A parametric dependent study is crucial for the accurate determination of transport coefficients such as shear viscosity. In this study, we calculate the shear viscosity of extended simple point charge water using a transverse current auto-correlation function (TCAF) from equilibrium molecular dynamics (EMD) and the periodic perturbation method from non-equilibrium molecular dynamics (NEMD) simulations for varying coupling time and system sizes. Results show that the shear viscosity calculated using EMD simulations with different thermostats varies significantly with coupling times and system size. The use of Berendsen and velocity-rescale thermostats in NEMD simulations generates a significant drift from the target temperature and results in an inconsistent shear viscosity with coupling time and system size. The use of Nosé–Hoover thermostat in NEMD simulations offers thermodynamic stability which results in a consistent shear viscosity for various coupling times and system sizes.     

High-speed sensing of microliter-order whole-blood viscosity using laser-induced capillary wave

The present paper introduces an innovative contact-free optical viscosity measurement technique, laser-induced capillary wave (LiCW) using pulsed YAG laser as a heating source, to measure whole-blood viscosity with only a microliter-order sample volume and measurement time on the millisecond order. In this method, interfering pulsed laser beams heat a whole-blood sample and generate a capillary wave, the amplitude of which is less than 10 nm with wavelength of 80–100 μm in the present experiment, caused by a spatially sinusoidal temperature distribution. The damped oscillation of the capillary wave, which is detected by a diffracted probing laser beam at the heated area, provides information regarding the viscosity and surface tension of the whole blood. To demonstrate the validity of the present laser-induced capillary wave viscometer, the viscosity of human whole blood taken from two healthy donors having different hematocrit values was measured using 90 μl sample volumes at 37°C. To consider the feasibility of the present technique for blood rheological studies, we discuss the characteristics of LiCW regarding the non-Newtonian behavior of blood, the velocity boundary layer, the existence of a free surface, and the temperature increase of the blood, and also demonstrate the capability of the method to sense the temporal evolution of blood viscosity with sampling frequency of 0.25 Hz.     

The failure of simple empirical relationships to predict the viscosity of mixed aqueous solutions of guanidine hydrochloride and glucose has important implications for the study of protein folding

Viscosities of aqueous solutions of guanidine hydrochloride (GuHCl) were measured in the presence of varying amounts of glucose. At high concentrations of glucose or GuHCl, the measured viscosities showed significant deviation from the values computed using a method proposed by Tanford (1966, J Biol Chem 241:3228-3232). This method was originally derived to allow the calculation of the effects of buffer or low concentrations of salts and other additives on the density and viscosity of aqueous solutions of urea or GuHCl. Recently it has been used to estimate the viscosity of denaturant solutions that contain high concentrations of viscogens. Our results show that the extrapolation of this approach to solutions of highly concentrated viscous co-solutes leads to significant errors. The implications for experimental studies of the viscosity dependence of conformational transitions in proteins is discussed.     

Effects of the non-Newtonian viscosity of blood on flows in a diseased arterial vessel. Part 1: Steady flows

Effects of the non-Newtonian viscosity of blood on a flow in a coronary arterial casting of man were studied numerically using a finite element method. Various constitutive models were examined to model the non-Newtonian viscosity of blood and their model constants were summarized. A method to incorporate the non-Newtonian viscosity of blood was introduced so that the viscosity could be calculated locally. The pressure drop, wall shear stress and velocity profiles for the case of blood viscosity were compared for the case of Newtonian viscosity (0.0345 poise). The effect of the non-Newtonian viscosity of blood on the overall pressure drop across the arterial casting was found to be significant at a flow of the Reynolds number of 100 or less. Also in the region of flow separation or recirculation, the non-Newtonian viscosity of blood yields larger wall shear stress than the Newtonian case. The origin of the non-Newtonian viscosity of blood was discussed in relation to the viscoelasticity and yield stress of blood.     

The effect of temperature on ATP levels in cells of an ascitic hepatoma

The effect of temperature on intracellular ATP levels in cells of the ascitic Hepatoma 129 has been determined using an UV-enzymic method. ATP levels are found to vary inversely with temperature over the range (4°–37°C) studied, and this variation is found to be reversible. An indication of a possible direct correlation of ADP level with temperature was also obtained. A correlation is deduced between rise in intracellular ATP level and decrease in cytoplasmic viscosity, both from the present results, and from similar effects recently reported by Landau ('61) for increase in pressure. The significance of these observations is discussed with reference to the model for cytoplasmic viscosity proposed by Marsland ('56) and the wider biological significance of this effect is explored.     

Intrinsic viscosity of bead models for macromolecules and bioparticles

A new method based on the fractal dimension dependence of the hydrodynamic radius is proposed for calculation of the intrinsic viscosity of bead models. The method describes properly the viscosity increment except for elongated structures such as linear aggregates and ellipsoids. It is expected to be useful for very compact structures, for which the volume correction does not improve the results calculated by the modified Oseen tensor. The results obtained for the viscosity increment lie between the volume corrected ones and those determined by the cubic substitution procedure. They are close to the values recalculated from the falling velocities of the models analyzed.     

Synthesis of oxidized guar gum by dry method and its application in reactive dye printing

The aim of this study was to prepare oxidized guar gum with a simple dry method, basing on guar gum, hydrogen peroxide and a small amount of solvent. To obtain a product with suitable viscosity for reactive dye printing, the effects of various factors such as the amount of oxidant and solvent, reaction temperature and time were studied with respect to the viscosity of reaction products. The product was characterized by Fourier transform infrared spectroscopy, size exclusion chromatography, scanning electron microscopy and differential scanning calorimetry. The hydrated rate of guar gum and oxidized guar gum was estimated through measuring the required time when their solutions (1%, w/v) reached the maximum viscosity. The effects of the salt concentration and pH on viscosity of the resultant product were studied. The mixed paste containing oxidized guar gum and carboxymethyl starch was prepared and its viscosity was determined by the viscometer. The rheological property of the mixed paste was appraised by the printing viscosity index. In addition, the applied effect of mixed paste in reactive dye printing was examined by assessing the fabric stiffness, color yield and sharp edge to the printed image in comparison with sodium alginate. And the results indicated that the mixed paste could partially replace sodium alginate as thickener in reactive dye printing. The study also showed that the method was low cost and eco-friendly and the product would have an extensive application in reactive dye printing.     

A new analysis method for the membrane viscosity from steady-state fluorescence depolarization

The absolute value of the viscosity in membrane lipid bilayers, which is different from the microviscosity advocated by Shinitzky, could be calculated from steady-state fluorescence depolarization of a hydrocarbon fluorophore, 1,6-diphenyl-1,3,5-hexatriene (DPH). This method was based on the theory of time-resolved fluorescence anisotropy and empirical relationships between fluorescence life time and the anisotropy parameters such as half cone angle in wobbling motion and wobbling diffusion rate of the fluorescent probe. Obtained viscosity values of various membranes from this method were consistent with those from time resolved method within experimental error.     

A vesicle microrheometer for high-throughput viscosity measurements of lipid and polymer membranes

Viscosity is a key property of cell membranes that controls mobility of embedded proteins and membrane remodeling. Measuring it is challenging because existing approaches involve complex experimental designs and/or models, and the applicability of some methods is limited to specific systems and membrane compositions. As a result there is scarcity of systematic data, and the reported values for membrane viscosity vary by orders of magnitude for the same system. Here, we show how viscosity of membranes can be easily obtained from the transient deformation of giant unilamellar vesicles. The approach enables a noninvasive, probe-independent, and high-throughput measurement of the viscosity of membranes made of lipids or polymers with a wide range of compositions and phase state. Using this novel method, we have collected a significant amount of data that provides insights into the relation between membrane viscosity, composition, and structure.     

Continuous capture of recombinant antibodies by ZnCl2 precipitation without polyethylene glycol

The capture of recombinant antibodies from cell culture broth is the first critical step of downstream processing. We were able to develop a precipitation‐based method for the capture and purification of monoclonal antibodies based on divalent cations, namely ZnCl₂. Traditional precipitation processes have to deal with high dilution factors especially for resolubilization and higher viscosity due to the use of PEG as precipitation or co‐precipitation agent. By the use of the crosslinking nature of divalent cations without the use of PEG, we kept viscosity from the supernatant and resolubilization dilution factors very low. This is especially beneficial for the solid–liquid separation for the harvest and wash of the precipitate in continuous mode. For this harvest and wash, we used tangential flow filtration that benefits a lot from low viscosity solutions, which minimizes the membrane fouling. With this precipitation based on ZnCl_2, we were able to implement a very lean and efficient process. We demonstrated precipitation studies with three different antibodies, Adalimumab, Trastuzumab, and Denosumab, and a continuous capture case study using tangential flow filtration for precipitate recovery. In this study, we achieved yields of 70%.     

Computer simulation of flagellar movement. IV. Properties of an oscillatory two-state cross-bridge model.

A stochastic computational method is used to examine the properties of a simple two-state cross-bridge model, of a type which has been shown previously to self-oscillate without requiring any feedback control of the active process. The force transients obtained with this model show the major features observed with oscillatory insect fibrillar flight muscle. The effects of viscosity and cross-bridge detachment rate on the frequency of oscillation of the model resemble the effects of viscosity and ATP concentration on flagellar oscillation, and the relationship between turnover rate and frequency of oscillation is also consistent with observations on flagella. However, the amplitude of oscillation of the model does not show the degree of frequency-independence which is typical of flagella.     

A comparison of five methods for obtaining the intrinsic viscosity of bovine serum albumin

Intrinsic viscosity [η] is a characteristic of proteins and other molecules related directly to their ability to disturb flow and indirectly to their size and shape. It is usually determined by extrapolating reduced viscosity to zero concentration. Four other methods for deriving [η] have been utilized by previous investigators. Studies of the intrinsic viscosity of bovine serum albumin had been carried out two years apart as a test of viscometry technique; the data obtained were used to compare the five methods. Four of the five produced [η] values ranging from 3.92 to 4.21 ml/g. Agreement was good between the two studies. The five methods were compared to each other using linearity of regression, statistical error of determination, effect of varying solvent time, and result obtained in different concentration ranges. By these four criteria, use of the regression of specific fluidity (1 ? 1/η_rel) with concentration was found superior to other methods. Its only deficiency was a requirement that solution density be corrected for at each concentration studied rather than applying a single correction for density after using kinematic viscosity data. All methods for deriving intrinsic viscosity are based on one of three equations; flow is expressed either in terms of reduced viscosity (Huggins), inherent viscosity (Kraemer), or specific fluidity. Of these three equations, specific fluidity is the most closely related both to theoretical analyses and to experimental studies of rigid spheres. There is abundant evidence in past reports that in contrast to rigid spheres, flexible polymers do not produce a linear rise in specific fluidity as their concentration increases, strongly suggesting that their molecular conformation is changing with concentration. A linear relation between fluidity and concentration has been observed for almost all proteins and protein mixtures studied. When this linear relation is present it indicates both that molecular conformation during flow is independent of concentration in the range studied and that the specific fluidity method for deriving intrinsic viscosity is the most appropriate.     

Estimating intercellular surface tension by laser-induced cell fusion

Intercellular surface tension is a key variable in understanding cellular mechanics. However, conventional methods are not well suited for measuring the absolute magnitude of intercellular surface tension because these methods require determination of the effective viscosity of the whole cell, a quantity that is difficult to measure. In this study, we present a novel method for estimating the intercellular surface tension at single-cell resolution. This method exploits the cytoplasmic flow that accompanies laser-induced cell fusion when the pressure difference between cells is large. Because the cytoplasmic viscosity can be measured using well-established technology, this method can be used to estimate the absolute magnitudes of tension. We applied this method to two-cell-stage embryos of the nematode Caenorhabditis elegans and estimated the intercellular surface tension to be in the 30-90 μN m(-1) range. Our estimate was in close agreement with cell-medium surface tensions measured at single-cell resolution.     

The apparent viscosity of the protoplasm of subepidermal stem basis cells of Pisum sativum. Relation to aging, drought tolerance and water stress

Plants of Pisum sativum L. cv. Alaska wilt resistant were subjected to two different water stress regimes under controlled environment conditions: watering was stopped either on the 7th day (early stress) or on the 21st day (late stress) after planting. Plants under the early stress regime developed drought tolerance (adapted), while those under late stress did not. The apparent viscosity of the protoplasm of subepidermal stem basis cells was analyzed by the centrifugation and plasmolysis form method during the entire growth period.
The apparent viscosity of the subepidermal stem basis cells changed with plant age and was highest in 3-week-old plants. In controls the relation of apparent viscosity to age was the same when measured under full turgor and in relaxed state. Under early stress condition, however, the pattern of the viscosity changes with plant age was significantly different for turgescent and relaxed cells. In four week old plants, a higher apparent viscosity was measured in relaxed adapted cells than in relaxed control cells. It is suggested that the higher apparent viscosity is the result of a delayed cell aging.
Apparent viscosity was inversely proportional to soil moisture content and the osmotic potential of the cell sap for the cells of late stress plants, whereas no clear relation was found for the cells of early stress plants. This difference may indicate two mechanisms of viscosity changes: 1) osmotic dehydration of the protoplasm under water stress (passive viscosity change), 2) changes in the amount, hydration or architecture of macromolecules present in the cytoplasm (active viscosity change). Whereas differences in the apparent viscosity between control and stressed cells may not be the cause of drought tolerance, they seem to indicate the development of drought tolerance. Water stress history and plant age were the most critical factors controlling the apparent viscosity changes observed.