•MicroScale Thermophoresis (MST) measures dissociation constants down to 1 picomolar.•MST can dissect different binding mechanism by distinct thermophoresis signals.•MST quantifies dissociation ...constants in 100% cell lysate.•MST can be used to determine thermodynamic parameters for interactions.•MST can be used to monitor enzyme kinetics and protein folding.
MicroScale Thermophoresis (MST) is a powerful technique to quantify biomolecular interactions. It is based on thermophoresis, the directed movement of molecules in a temperature gradient, which strongly depends on a variety of molecular properties such as size, charge, hydration shell or conformation. Thus, this technique is highly sensitive to virtually any change in molecular properties, allowing for a precise quantification of molecular events independent of the size or nature of the investigated specimen.
During a MST experiment, a temperature gradient is induced by an infrared laser. The directed movement of molecules through the temperature gradient is detected and quantified using either covalently attached or intrinsic fluorophores. By combining the precision of fluorescence detection with the variability and sensitivity of thermophoresis, MST provides a flexible, robust and fast way to dissect molecular interactions.
In this review, we present recent progress and developments in MST technology and focus on MST applications beyond standard biomolecular interaction studies. By using different model systems, we introduce alternative MST applications – such as determination of binding stoichiometries and binding modes, analysis of protein unfolding, thermodynamics and enzyme kinetics. In addition, wedemonstrate the capability of MST to quantify high-affinity interactions with dissociation constants (Kds) in the low picomolar (pM) range as well as protein–protein interactions in pure mammalian cell lysates.
In biomass and waste combustion, the release of KCl into the gas phase results in its deposition on heat exchanger surfaces, enhancing their stickiness and accelerating deposit buildup from incoming ...ash particles. Despite this, the mechanisms governing KCl deposition remain inadequately explored. KCl has been observed to form either a fine, powdery layer through thermophoresis of nucleated KCl particles or a crystal-like, coarse layer through heterogeneous condensation of KCl vapor. However, the quantitative assessment of coarse and fine layer formation has never been performed. In this study, an entrained flow reactor and mechanistic model were employed to analyze KCl deposition behavior, examining the effect of probe temperature and exposure time. Quantification of both the coarse and fine deposit layers was accomplished. Probe temperature was found to have no influence on the deposition in the range from 300 °C – 595 °C. For coarse layer formation, the critical factor was the development of a fine layer upon which the crystal-like structure of the coarse layer grows. Lower probe temperatures accelerate initial layer formation, initiating the coarse section sooner. Additionally, it slightly increases nucleation and condensation, however, nucleation is increased stronger resulting in a lower ratio of coarse to fine deposit. The quantification of the coarse mass, in conjunction with the simulation, provides a first estimation of the critical mass required to initiate heterogeneous condensation on a steel tube. In this study, a critical mass of 1.26e-3 g/cm2 was determined.
The present study aims to investigate the impact of nanoparticle migration due to Brownian motion and thermophoresis on Ag-MgO/Water hybrid nanofluid natural convection. An enclosure with sinusoidal ...wavy walls is considered for this investigation; right and cold walls of this enclosure are in constant temperature while the upper and bottom walls are insulated. This simulation follows Buongiorno's mathematical model; Brownian and thermophoresis diffusion of Ag occurs in MgO-Water nanofluid while the diffusion of MgO happens in Ag-water nanofluid. The result indicates that Nu number increments up to 11% by increasing thermophoresis diffusion for both nanoparticles. Also, increasing Brownian diffusion of Ag augments nanoparticle concentration on hot wall, while the accumulation of nanoparticles unifies by incrementing Brownian diffusion of MgO.
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•Ag − MgO/water hybrid nanofluid utilizing Buongiorno's mathematical model.•Nanoparticle accumulates better on the cold wall due to thermophoresis migration.•Nu number increments up to 11% by increasing thermophoresis diffusion.•The accumulation of nanoparticles unifies by augmenting DBMgO.
•Heat transport and entropy generation of thermodynamic second law is considered.•A mathematical model is presented for entropy generation.•Fluid is conducting electrically through applied magnetic ...flux.•Optimal homotopy technique is employed to tackle the simplified equations.
Flow of Williamson nanofluid over a stretching sheet is addressed. Entropy generation is modeled by second thermodynamics law. Present model examines the momentum, heat, mass and entropy generation. Joule heating, viscous dissipation and chemical reaction are considered in the modeling. Fluid is conducting electrically through applied magnetic field. Optimal homotopy analysis method is implemented for the solutions. Series solutions convergence by residual errors is ensured. Outcomes of magnetic parameter. Prandtl number, Brownian motion, chemical reaction parameter, thermophoresis and Brinkman number are examined. Moreover coefficient of skin friction and heat transfer rate (Nusselt number) are computed and examined. Heat transfer rate by Brownian parameter is increased. Furthermore entropy generation rate increases for higher magnetic parameter.
In this study, two dimensional mixed convective MHD stagnation point flow of Casson fluid past an infinite plate in porous medium is considered. Effects of thermophoresis, Brownian motion, non-linear ...thermal radiation, heat generation and chemical reaction are considered. The governing differential equations are modified using similarity transformation. System of ODE is solved using HAM. Effects of different vital parameters on velocity, temperature and concentration profiles are shown graphically. The numerical values of the velocity, temperature and concentration gradient are also obtained in tabular form. It is illustrated that, mixed convection parameter λ, buoyancy force parameter N, non-linear thermal radiation parameter Nr and heat generation parameter H tends to improve motion of the fluid throughout the flow field. It is also seen that, heat and mass transfer process improve with Brownian motion Nb and thermophoresis parameter Nt. Brownian motion and thermophoresis parameters have negative impacts on temperature gradient.
Targeting RNA with small molecules is an emerging field. While several ligands for different RNA targets are reported, structure-based virtual screenings (VSs) against RNAs are still rare. Here, we ...elucidated the general capabilities of protein-based docking programs to reproduce native binding modes of small-molecule RNA ligands and to discriminate known binders from decoys by the scoring function. The programs were found to perform similar compared to the RNA-based docking tool rDOCK, and the challenges faced during docking, namely, protomer and tautomer selection, target dynamics, and explicit solvent, do not largely differ from challenges in conventional protein-ligand docking. A prospective VS with the Bacillus subtilis preQ1-riboswitch aptamer domain performed with FRED, HYBRID, and FlexX followed by microscale thermophoresis assays identified six active compounds out of 23 tested VS hits with potencies between 29.5 nM and 11.0 μM. The hits were selected not solely based on their docking score but for resembling key interactions of the native ligand. Therefore, this study demonstrates the general feasibility to perform structure-based VSs against RNA targets, while at the same time it highlights pitfalls and their potential solutions when executing RNA–ligand docking.
With rising popularity of microscale thermophoresis for the characterisation of protein-ligand binding reactions and possible applications in microfluidic devices, there is a growing interest in ...considering thermodiffusion in the context of life sciences. But although the understanding of thermodiffusion in non-polar mixtures has grown rapidly in recent years, predictions for associated mixtures like aqueous solutions remain challenging. This review aims to give an overview of the literature on thermodiffusion in aqueous systems, show the difficulties in theoretical description that arise from the non-ideal behaviour of water-mixtures, and highlight the relevance of thermodiffusion in a biological context. We find that the thermodiffusion in aqueous systems is dominated by contributions from heat of transfer, hydrogen bond interactions and charge effects. However, the separation of these effects is often difficult, especially in case of biological systems where a systematic exclusion of contributions may not be feasible.
Intention here is to discuss the physical features of MHD stagnation point flow of Williamson nanomaterial over a stretched surface. Cattaneo-Christov Double Diffusion (CCDD) concept is incorporated ...to deliberate the behaviors of mass and heat transfer rates. These models are based upon Fick's and Fourier's laws versus solutal and thermal relaxation times. Innovative characteristics regarding thermophoresis and Brownian diffusion effect are also scrutinized. Relevant transformations are used to reduce the nonlinear expression to ordinary one. Convergent solution by optimal homotopy analysis technique (OHAM) is constructed. The average residual error is computed through OHAM Discussion is arranged for velocity, concentration and temperature. Skin friction co-efficient, temperature gradient and mass transfer rate are graphically discussed. Velocity boosts up significantly via velocity ratio parameter. Temperature and velocity have opposite effects for Weissenberg number. Concentration of the material particles increases against larger estimations of thermal relaxation time.
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•A chimeric aptamer of AFB1 firstly engineered by a truncation and splicing-based post-selection strategy.•MST proved the chimeric aptamer had 188-fold higher affinity for AFB1 than ...the originally-selected aptamer.•ITC and molecular docking revealed the dual binding sites of the chimeric aptamer.•A chimeric aptamer-based AuNPs aptasensor achieved superior sensitivity, specificity, and repeatability for real samples.•The proof-of concept can be referred for other aptamer-related studies.
In this study, a bivalent anti-Aflatoxin B1 (AFB1) chimeric aptamer (B72) was firstly engineered by a rational truncation and splicing-based post-selection strategy. Compared with that of the originally-selected anti-AFB1 aptamer (B50), a 188-fold enhanced affinity of B72 was revealed by microscale thermophoresis (MST) assay. In addition, isothermal titration calorimetry (ITC) research and molecular docking simulation affirmed consistently that B72 possessed dual binding sites for AFB1. A gold nanoparticles (AuNPs) aptasensor was developed to further validate the real-world applicability of the chimeric aptamer by colorimetric assay of AFB1 based on the peroxidase-like activity of AuNPs to catalyze 3,3,5,5-tetramethylbenzidine (TMB) with the assistance of H2O2. The aptasensor could identify AFB1 in corn oil with a limit of detection (LOD) of 1.88 nM and a linear range of 5–5120 nM. Moreover, good recovery rates of 91.5–117.6% and low relative standard deviations below 9% were obtained, suggesting the good accuracy of the developed aptasensor. The study could thus be referred as a universal post-selection strategy for engineering high affinity aptamers and constructing novel aptamer-based biosensing platforms for highly sensitive and specific analysis of other targets.
Here, the synthesis of linear polyglycerols bearing multiple copies of mono and dimannosides (LPG40Man0.60 and LPG40(Manα1,2Man)0.60) is demonstrated. A method based on label‐free microscale ...thermophoresis is optimized to determine the direct binding affinity of multivalent mannosides for Escherichia coli (E. coli) strain ORN178 that produces the fimbriae protein FimH. It is observed that the LPG40(Manα1,2Man)0.60 exhibits only a modest onefold improvement in binding as compared to LPG40Man0.60. Nevertheless, both the multivalent mannosides display remarkably very low nm binding constant (Kd) in contrast to the high µm Kd of the single α‐d‐methylmannoside for intact E. coli ORN178 particles. Furthermore, in an adhesion‐inhibition assay, both multivalent mannosides show 50% inhibition of bacteria adhesion to the HT‐29 colon cells at low µm concentrations.
The multivalent mono and dimannosides based on linear polyglycerol identical in size and glycan density are synthesized. The label‐free microscale thermophoresis is employed to determine the binding affinities of multivalent scaffolds against whole bacteria. Both multivalent mannosides display remarkably very low nm binding constant (Kd) in contrast to very high µm Kd of α‐d‐methylmannoside for intact Escherichia coli.
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