The substrate-binding domains of the osmoregulatory ABC importer OpuA transiently interact.

Bacteria utilize various strategies to prevent internal dehydration during hypertonic stress. A common approach to countering the effects of the stress is to import compatible solutes such as glycine betaine, leading to simultaneous passive water fluxes following the osmotic gradient. OpuA from Lactococcus lactis is a type I ABC-importer that uses two substrate-binding domains (SBDs) to capture extracellular glycine betaine and deliver the substrate to the transmembrane domains for subsequent transport. OpuA senses osmotic stress via changes in the internal ionic strength and is furthermore regulated by the 2nd messenger cyclic-di-AMP. We now show, by means of solution-based single-molecule FRET and analysis with multi-parameter photon-by-photon hidden Markov modeling, that the SBDs transiently interact in an ionic strength-dependent manner. The smFRET data are in accordance with the apparent cooperativity in transport and supported by new cryo-EM data of OpuA. We propose that the physical interactions between SBDs and cooperativity in substrate delivery are part of the transport mechanism.

NAGase sensing in 3% milk: FET-based specific and label-free sensing in ultra-small samples of high ionic strength and high concentration of non-specific proteins.

Biosensing with biological field-effect transistors (bioFETs) is a promising technology toward specific, label-free, and multiplexed sensing in ultra-small samples. The current study employs the field-effect meta-nano-channel biosensor (MNC biosensor) for the detection of the enzyme N-acetyl-beta-D-glucosaminidase (NAGase), a biomarker for milk cow infections. The measurements are performed in a 0.5 µL drops of 3% commercial milk spiked with NAGase concentrations in the range of 30.3 aM-3.03 µM (Note that there is no background NAGase concentration in commercial milk). Specific and label-free sensing of NAGase is demonstrated with a limit-of-detection of 30.3 aM, a dynamic range of 11 orders of magnitude and with excellent linearity and sensitivity. Additional two important research outcomes are reported. First, the ionic strength of the examined milk is ∼120 mM which implies a bulk Debye screening length <1 nm. Conventionally, a 1 nm Debye length excludes the possibility of sensing with a recognition layer composed of surface bound anti-NAGase antibodies with a size of ∼10 nm. This apparent contradiction is removed considering the ample literature reporting antibody adsorption in a predominantly surface tilted configuration (side-on, flat-on, etc.). Secondly, milk contains a non-specific background protein concentration of 33 mg/ml, in addition to considerable amounts of micron-size heterogeneous fat structures. The reported sensing was performed without the customarily exercised surface blocking and without washing of the non-specific signal. This suggests that the role of non-specific adsorption to the BioFET sensing signal needs to be further evaluated. Control measurements are reported.

On the covalent nature of lysine polyphosphorylation.

Post-translational modifications of proteins (PTMs) introduce an extra layer of complexity to cellular regulation. Although phosphorylation of serine, threonine, and tyrosine residues is well-known as PTMs, lysine is, in fact, the most heavily modified amino acid, with over 30 types of PTMs on lysine having been characterized. One of the most recently discovered PTMs on lysine residues is polyphosphorylation, which sees linear chains of inorganic polyphosphates (polyP) attached to lysine residues. The labile nature of phosphoramidate bonds raises the question of whether this modification is covalent in nature. Here, we used buffers with very high ionic strength, which would disrupt any non-covalent interactions, and confirmed that lysine polyphosphorylation occurs covalently on proteins containing PASK domains (polyacidic, serine-, and lysine-rich), such as the budding yeast protein nuclear signal recognition 1 (Nsr1) and the mammalian protein nucleolin. This Matters Arising Response paper addresses the Neville et al. (2024) Matters Arising paper, published concurrently in Molecular Cell.

Characterization of Freezing Processes in Drug Substance Bottles by Ice Core Sampling.

Freezing of biological drug substance (DS) is a critical unit operation that may impact product quality, potentially leading to protein aggregation and sub-visible particle formation. Cryo-concentration has been identified as a critical parameter to impact protein stability during freezing and should therefore be minimized. The macroscopic cryo-concentration, in the following only referred to as cryo-concentration, is majorly influenced by the freezing rate, which is in turn impacted by product independent process parameters such as the DS container, its size and fill level, and the freezing equipment. (At-scale) process characterization studies are crucial to understand and optimize freezing processes. However, evaluating cryo-concentration requires sampling of the frozen bulk, which is typically performed by cutting the ice block into pieces for subsequent analysis. Also, the large amount of product requirement for these studies is a major limitation. In this study, we report the development of a simple methodology for experimental characterization of frozen DS in bottles at relevant scale using a surrogate solution. The novel ice core sampling technique identifies the axial ice core in the center to be indicative for cryo-concentration, which was measured by osmolality, and concentrations of histidine and polysorbate 80 (PS80), whereas osmolality revealed to be a sensitive read-out. Finally, we exemplify the suitability of the method to study cryo-concentration in DS bottles by comparing cryo-concentrations from different freezing protocols (-80°C vs -40°C). Prolonged stress times during freezing correlated to a higher extent of cryo-concentration quantified by osmolality in the axial center of a 2 L DS bottle.

Efficacy of hydroxypropyl-guar drops in improving tear film index and ocular surface dynamics using two treatment methods under a controlled desiccating environment.

AIM: This study aimed to assess the efficacy of hp-guar eye drops on tear film index and ocular surface dynamics under desiccating conditions using protection and relief treatment modalities. METHODOLOGY: The 12 normal, non-dry eye participants were subjected to adverse environmental conditions using a Controlled Environment Chamber (CEC) where the relative humidity (RH) was 5% and the ambient temperature was 21 °C. The participants were screened for ocular symptoms, tear osmolarity, ocular surface temperature (OST), tear production using the Ocular Surface Disease Index questionnaire (OSDI), OcuSense TearLab Osmometer, FLIR System ThermaCAM P620, and Schirmer strips. Tear production was calculated by the Tear Function Index test (TFI). RESULTS: The mean tear film osmolarity decreased significantly from 296 mOsm/L at 40% RH to 285 mOsm/L at 5% RH (p = 0.01). Conflicting responses were seen for osmolarity in protection and relief. Mean tear osmolarity was significantly higher in the protection method in comparison to the relief method (p = 0.005). The mean TFI increased from 557 at 40% to 854 at 5% (p = 0.02). A significant increase in TFI was observed in the relief method in comparison with both 40% (p = 0.001) and 5% (p = 0.04). In the relief method, the mean TFI score went up to 1139 when hp-guar was installed. A significant improvement in ocular comfort was experienced in both the protection (p = 0.041) and relief (p = 0.010) methods at 5% RH. The instillation of hp-guar drops in the relief method resulted in a significant reduction in OST. The mean OST dropped to 33.01 ºC, significantly lower than the recorded OST for both normal (p = 0.040) and dry (p = 0.014) environmental conditions. CONCLUSION: Hp-guar drops significantly improve tear film parameters under a desiccating environment, however, tear film parameters respond differently to the management modalities. In the protection method, tear film osmolarity was protected against a dry environment, while in the relief mode, an improvement in tear production and a decrease in ocular surface temperature were seen. Hp-guar performance could be maximized for the management of exposure to adverse environments by using a treatment protocol that targets the most affected parameters in each group of patients. Using CEC has the potential to provide researchers with a readily available method to evaluate the efficiency of tear supplementation.

Change in Osmotic Pressure Influences the Absorption Spectrum of Hemoglobin inside Red Blood Cells.

Absorption spectra of red blood cell (RBC) suspensions are investigated in an osmolarity range in the medium from 200 mOsm to 900 mOsm. Three spectral parameters are used to characterize the process of swelling or shrinkage of RBC-the absorbance at 700 nm, the Soret peak height relative to the spectrum background, and the Soret peak wavelength. We show that with an increase in the osmolarity, the absorbance at 700 nm increases and the Soret peak relative height decreases. These changes are related to the changes in the RBC volume and the resulting increase in the hemoglobin intracellular concentration and index of refraction. Confocal microscopy and flow cytometry measurements supported these conclusions. The maximum wavelength of the Soret peak increases with increasing osmolarity due to changes in the oxygenation state of hemoglobin. Using these spectrum parameters, the process of osmosis in RBCs can be followed in real time, but it can also be applied to various processes, leading to changes in the volume and shape of RBCs. Therefore, we conclude that UV-Vis absorption spectrophotometry offers a convenient, easily accessible, and cost-effective method to monitor changes in RBC, which can find applications in the field of drug discovery and diagnostics of RBC and hemoglobin disorders.

Adsorption-desorption of 241Am(â ¢) on montmorillonite colloids and quartz sand: Effects of pH, ionic strength, colloid concentration and grain size.

Clay colloids in the subsurface environment have a strong adsorption capacity for radionuclides, and the mobile colloids will carry the nuclides for migration, which would promote the movability of radionuclides in the groundwater environment and pose a threat to the ecosphere. The investigations of the adsorption/desorption behaviors of radionuclides in colloids and porous media are significant for the evaluation of the geological disposal of radioactive wastes. To illustrate the adsorption/desorption behaviors of 241Am(Ⅲ) in Na-montmorillonite colloid and/or quartz sand systems at different pH (5, 7 and 9), ionic strengths (0, 0.1 and 5 mM), colloid concentrations (300 and 900 mg/L), nuclide concentrations (500, 800, 1100 and 1400 Bq/mL) and grain sizes (40 and 60 mesh), a series of batch sorption-desorption experiments were conducted. Combining the analysis of the physical and chemical properties of Na-montmorillonite with the Freundlich model, the influencing mechanism of different controlling factors is discussed. The experimental results show that the adsorption/desorption behaviors of 241Am(Ⅲ) in Na-montmorillonite colloid and/or quartz sand strongly are influenced by the pH value and ionic strength of a solution, the colloid concentration as well as quartz sand grain size. The adsorption and desorption isotherms within all the experimental conditions could be well-fitted by the Freundlich model and the correlation coefficients (R2) are bigger than 0.9. With the increase in pH, the adsorption partition coefficient (Kd) at 241Am(Ⅲ)-Na-montmorillonite colloid two-phase system and 241Am(Ⅲ)-Na-montmorillonite colloid-quartz sand three-phase system presents a trend which increases firstly followed by decreasing, due to the changes in the morphology of Am with pH. The Kd of 241Am(Ⅲ) adsorption on montmorillonite colloid and quartz sand decreases with increasing in ionic strength, which is mainly attributed to the competitive adsorption, surface complexation and the reduction of surface zeta potential. Additionally, the Kd increases with increasing colloid concentrations because of the increase in adsorption sites. When the mean grain diameter changes from 0.45 to 0.3 mm, the adsorption variation trends of 241Am(Ⅲ) remain basically unchanged. The research results obtained in this work are meaningful and helpful in understanding the migration behaviors of radionuclides in the underground environment.

Single-Molecule Diffusivity Quantification Unveils Ubiquitous Net Charge-Driven Protein-Protein Interaction.

Recent microscopy and nuclear magnetic resonance (NMR) studies have noticed substantial suppression of intracellular diffusion for positively charged proteins, suggesting an overlooked role of electrostatic attraction in nonspecific protein interactions in a predominantly negatively charged intracellular environment. Utilizing single-molecule detection and statistics, here, we quantify in aqueous solutions how protein diffusion, in the limit of low diffuser concentration to avoid aggregate/coacervate formation, is modulated by differently charged interactor proteins over wide concentration ranges. We thus report substantially suppressed diffusion when oppositely charged interactors are added at parts per million levels, yet unvaried diffusivities when same-charge interactors are added beyond 1%. The electrostatic attraction-driven suppression of diffusion is sensitive to the protein net charge states, as probed by varying the solution pH and ionic strength or chemically modifying the proteins and is robust across different diffuser-interactor pairs. By converting the measured diffusivities to diffuser diameters, we further show that in the limit of excess interactors, a positively charged diffuser molecule effectively drags along just one monolayer of negatively charged interactors, where further interactions stop. We thus unveil ubiquitous, net charge-driven protein-protein interactions and shed new light on the mechanism of charge-based diffusion suppression in living cells.

Cotransport of aged biochar colloids and thallium(I) in water-saturated porous media: Impact of the ionic strength, pH and aging degree.

Biochar colloids entering the soil undergo aging over time and exhibit strong capabilities in adsorbing and transporting pollutants. Therefore, investigating the cotransport of aged biochar colloids and thallium (Tl(I)) in quartz sand media is crucial for understanding Tl(I) migration in underground environments. This study investigated the migration of biochar colloids with two different aging degrees and Tl(I) in quartz sand media at various pH and ionic strengths (ISs). The results revealed that under all ISs and pH, 30%AWB (biochar aged with 30 % (w/w) HNO3) inhibited Tl(I) migration in media. This inhibition primarily arose from the introduction of hydroxyl and carboxyl groups during aging, which significantly enhanced colloid adsorption onto Tl(I). At lower ISs, 30%AWB colloids exhibited greater inhibition of Tl(I) migration due to their increased adsorption capacity. Additionally, aging promoted the migration of biochar colloids in the media. Greater biochar aging notably enhanced this promotion, potentially owing to reduced colloidal particle size and the formation of biochar derivatives. Moreover, 50%AWB (biochar aged with 50 % (w/w) HNO3) inhibited Tl(I) migration under low ISs but had almost no impact under high ISs. Nonetheless, at high pH, 50%AWB colloids facilitated Tl(I) migration. This phenomenon might be attributed to the inhibitory effect of aged biochar colloids on Tl(I) adsorption onto media at a high pH, as well as the stable binding between Tl(I) and aged biochar colloids. This study discusses the cotransport of biochar with various degrees of aging and Tl(I) in media, providing insights into remediating soils contaminated with Tl.

Transport and retention of ciprofloxacin with presence of multi-walled carbon nanotubes in the saturated porous media: impacts of ionic strength and cation types.

The environmental fate and risks of ciprofloxacin (CIP) in the subsurface have raised intensive concerns. Herein, the transport behaviors of CIP in both saturated quartz sand and sand/multi-walled carbon nanotubes (MWCNTs) mixtures under different solution ionic strength of the solution and coexisting cation types were investigated. Batch adsorption experiments highlighted growing adsorptive capacity for CIP with the increasing content of MWCNTs in the MWCNTs-quartz sand mixtures (from 0.5% to 1.5%, w/w). Breakthrough curves (BTCs) of CIP in the MWCNTs-quartz sand mixtures were well fitted by the two-site chemical nonequilibrium model (R2 > 0.833). The estimated retardation factors for CIP increased from 9.68 to 282 with growing content of MWCNTs in the sand column, suggesting the presence of MWCNTs significantly inhibited the transport of CIP in saturated porous media. Moreover, the values of retardation factors are negatively correlated with the ionic strength and higher ionic strength could facilitate the transport of CIP in the saturated porous media. Compared with monovalent cations (Na+), the presence of divalent cations (Ca2+) significantly facilitated the transport of CIP in the columns due to the complexation between CIP and Ca2+ as well as deposition of MWCNTs aggregates on the sand surface. Results regarding CIP retention in columns indicated that MWCNTs could enhance the accumulation of CIP in the layers close to the influent of sand columns, while they could hinder upward transport of CIP to the effluent. This study improves our understanding for transport behaviors and environmental risk assessments of CIP in the saturated porous media with MWCNTs.

Compatibility studies of selected multichamber bag parenteral nutrition with fluconazole.

OBJECTIVE: Fluconazole (FLZ) is a drug widely used in the treatment of fungal infections including the treatment of immunocompromised patients, HIV-infected patients, and cancer patients. Critically ill patients often require the administration of drugs with parenteral nutrition (PN). The safety of this combination should be defined before the drug and PN are administered in one infusion line. This study aimed to determine the compatibility of FLZ with six selected multichamber bag parenteral nutrition. METHODS: FLZ solution for infusion was combined with PNs in appropriate proportions, considering most clinical situations resulting from different possible administration rates of the preparations. Samples were visually assessed, and pH, osmolality, turbidity, particle size (dynamic light scattering and light obscuration methods), and zeta potential were measured. These measurements were made immediately after combining the solutions and after 4 h of storage at 23 ± 1°C. RESULTS: FLZ combined with PNs did not cause changes observed visually. The turbidity of the samples was <0.4 NTU. The average particle size of the lipid emulsion was below 300 nm, and the PFAT5 parameter was ≤0.02%. The absolute value of the zeta potential of the PN + FLZ samples was higher for 5 out of 6 PN than the corresponding value for PN immediately after activation. Changes in pH and osmolality during 4 h of sample observations were within acceptable limits. CONCLUSION: Compatibility of the FLZ with six multichamber bag PN was confirmed. Hence, those preparations can be administered to patients in one infusion line using the Y-site.

Combining Scattering Experiments and Colloid Theory to Characterize Charge Effects in Concentrated Antibody Solutions.

Charges and their contribution to protein-protein interactions are essential for the key structural and dynamic properties of monoclonal antibody (mAb) solutions. In fact, they influence the apparent molecular weight, the static structure factor, the collective diffusion coefficient, or the relative viscosity, and their concentration dependence. Further, charges play an important role in the colloidal stability of mAbs. There exist standard experimental tools to characterize mAb net charges, such as the measurement of the electrophoretic mobility, the second virial coefficient, or the diffusion interaction parameter. However, the resulting values are difficult to directly relate to the actual overall net charge of the antibody and to theoretical predictions based on its known molecular structure. Here, we report the results of a systematic investigation of the solution properties of a charged IgG1 mAb as a function of concentration and ionic strength using a combination of electrophoretic measurements, static and dynamic light scattering, small-angle X-ray scattering, and tracer particle-based microrheology. We analyze and interpret the experimental results using established colloid theory and coarse-grained computer simulations. We discuss the potential and limits of colloidal models for the description of the interaction effects of charged mAbs, in particular pointing out the importance of incorporating shape and charge anisotropy when attempting to predict structural and dynamic solution properties at high concentrations.

Solution Ionic Strength Can Modulate Functional Loop Conformations in E. coli Dihydrofolate Reductase.

The observation of multiple conformations of a functional loop (termed M20) in the Escherichia coli dihydrofolate reductase (ecDHFR) enzyme triggered the proposition that large-scale motions of protein structural elements contribute to enzyme catalysis. The transition of the M20 loop from a closed conformation to an occluded conformation was thought to aid the rate-limiting release of the products. However, the influence of charged species in the solution environment on the observed M20 loop conformations, independent of charged ligands bound to the enzyme, had not been considered. Molecular dynamics simulations of ecDHFR in model CaCl2 solutions of varying molar ionic strengths IM reveal a substantial free energy barrier between occluded and closed M20 loop states at IM exceeding the E. coli threshold (∼0.24 M). This barrier may facilitate crystallization of ecDHFR in the occluded state, consistent with ecDHFR structures obtained at IM exceeding 0.3 M. At lower IM (≤0.15 M), the M20 loop can explore the occluded state, but prefers an open/partially closed conformation, again consistent with ecDHFR structures. Our findings caution against using ecDHFR structures obtained at nonphysiological ionic strengths in interpreting catalytic events or in structure-based drug design.

Mechanistic study of highly effective phosphate removal from aqueous solutions over a new lanthanum carbonate fabricated carbon nanotube film.

The effective purification of phosphate-containing wastewater is considered as increasingly important. In this study, a highly effective LC-CNT film was developed for efficient phosphate removal. Kinetic results showed that the adsorbent exhibited an improved mass transfer efficiency and a fast adsorption rate during adsorption (reaching 80% and 100% equilibrium adsorption capacity within 175 and 270 min, respectively). Kinetic model analysis suggested that the adsorption was a combined chemical physical process. Isotherm study revealed that the LC-CNT film showed a superior adsorption capacity (178.6 mg/g, estimated from the Langmuir model) with multiple adsorption mechanisms. pH study suggested that surface complexation and ligand exchange played important roles during adsorption, and the adsorbent worked well within the pH range of 3-7 with little La leakage. The ionic strength and competing anions showed little influence on the adsorbent effectiveness except for the carbonate and sulfate ions. The characterization and mechanism study revealed that the phosphate adsorption of the LC-CNT film was controlled by inner-sphere complexation, outer-sphere complexation and surface precipitation. The LC-CNT film also showed excellent regenerability and stability in cycling runs, further demonstrating its potential in industrial applications.

Rapid Enrichment of a Native Multipass Transmembrane Protein via Cell Membrane Electrophoresis through Buffer pH and Ionic Strength Adjustment.

Supported membrane electrophoresis is a promising technique for collecting membrane proteins in native bilayer environments. However, the slow mobility of typical transmembrane proteins has impeded the technique's advancement. Here, we successfully applied cell membrane electrophoresis to rapidly enrich a 12-transmembrane helix protein, glucose transporter 1 with antibodies (GLUT1 complex), by tuning the buffer pH and ionic strength. The identified conditions allowed the separation of the GLUT1 complex and a lipid probe, Fast-DiO, within a native-like environment in a few minutes. A force model was developed to account for distinct electric and drag forces acting on the transmembrane and aqueous-exposed portion of a transmembrane protein as well as the electroosmotic force. This model not only elucidates the impact of size and charge properties of transmembrane proteins but also highlights the influence of pH and ionic strength on the driving forces and, consequently, electrophoretic mobility. Model predictions align well with experimentally measured electrophoretic mobilities of the GLUT1 complex and Fast-DiO at various pH and ionic strengths as well as with several lipid probes, lipid-anchored proteins, and reconstituted membrane proteins from previous studies. Force analyses revealed the substantial membrane drag of the GLUT1 complex, significantly slowing down electrophoretic mobility. Besides, the counterbalance of similar magnitudes of electroosmotic and electric forces results in a small net driving force and, consequently, reduced mobility under typical neutral pH conditions. Our results further highlight how the size and charge properties of transmembrane proteins influence the suitable range of operating conditions for effective movement, providing potential applications for concentrating and isolating membrane proteins within this platform.

Refinement of a technique for collecting and evaluating the osmolality of haemolymph from Drosophila larvae.

Ex vivo physiological experiments using small insect models such as Drosophila larvae have become increasingly useful to address fundamental biological questions. To perform such experiments, various artificial saline solutions have been developed, but their osmolality varies significantly from one to the next. Such a variation of osmolality stems, in part, from the difficulty of determining the true value of haemolymph osmolality in Drosophila larvae. Thus, there is a pressing need to refine protocols for collecting and measuring the osmolality of the larval haemolymph. Two major obstacles are thought to impede the accurate analysis of haemolymph collected from small insects: melanin formation and gut-derived contamination. Here, we greatly refined existing haemolymph collection methods, evaluated the purity of the collected haemolymph under melanin-free conditions, and concluded that the true value of haemolymph osmolality is close to 306.0 mOsm kg-1 in Drosophila larvae.

The new preservative-free ophthalmic formulation of bilastine 0.6% preserves the ocular surface epithelial integrity in a comparative in vitro study.

Allergic conjunctivitis (AC) is the most common form of allergic eye disease and an increasingly prevalent condition. Topical eye drop treatments are the usual approach for managing AC, although their impact on the ocular surface is not frequently investigated. The aim of this study was to perform a comparative physicochemical characterization, and in vitro biological evaluations in primary conjunctival and corneal epithelial cells of the new multidose preservative-free bilastine 0.6% and main commercially available eye drops. MTT assay was used to measure cell viability; oxidative stress was analyzed with a ROS-sensitive probe; and apoptosis was evaluated monitoring caspase 3/7 activation. Differences in pH value, osmolarity, viscosity and phosphate levels were identified. Among all formulations, bilastine exhibited pH, osmolarity and viscosity values closer to tear film (7.4, 300 mOsm/l and ~ 1.5-10 mPa·s, respectively), and was the only phosphates-free solution. Single-dose ketotifen did not induce ROS production, and single-dose azelastine and bilastine only induced a mild increase. Bilastine and single-dose ketotifen and azelastine showed high survival rates attributable to the absence of preservative in its formulation, not inducing caspase-3/7-mediated apoptosis after 24 h. Our findings support the use of the new bilastine 0.6% for treating patients with AC to preserve and maintain the integrity of the ocular surface.

Rapid hydrolysis of NO2 at High Ionic Strengths of Deliquesced Aerosol Particles.

Nitrogen dioxide (NO2) hydrolysis in deliquesced aerosol particles forms nitrous acid and nitrate and thus impacts air quality, climate, and the nitrogen cycle. Traditionally, it is considered to proceed far too slowly in the atmosphere. However, the significance of this process is highly uncertain because kinetic studies have only been made in dilute aqueous solutions but not under high ionic strength conditions of the aerosol particles. Here, we use laboratory experiments, air quality models, and field measurements to examine the effect of the ionic strength on the reaction kinetics of NO2 hydrolysis. We find that high ionic strengths (I) enhance the reaction rate constants (kI) by more than an order of magnitude compared to that at infinite dilution (kI=0), yielding log10(kI/kI=0) = 0.04I or rate enhancement factor = 100.04I. A state-of-the-art air quality model shows that the enhanced NO2 hydrolysis reduces the negative bias in the simulated concentrations of nitrous acid by 28% on average when compared to field observations over the North China Plain. Rapid NO2 hydrolysis also enhances the levels of nitrous acid in other polluted regions such as North India and further promotes atmospheric oxidation capacity. This study highlights the need to evaluate various reaction kinetics of atmospheric aerosols with high ionic strengths.

The influence of charge on the translation of the sandwich ELISA approach to electronic biosensors.

The sandwich approach, whereby an antigen is captured by a primary antibody and detected by a secondary antibody, is commonly used to improve the selectivity and sensitivity of enzyme-linked immunosorbent assays (ELISA). This work details the experimental factors that impact the reliable translation of this sandwich approach to two commonly used electronic biosensors, namely potentiometric and impedimetric biosensors. Previous studies have demonstrated the Debye screening limitations associated with potentiometric biosensors. However, the correlation between the ionic strength of the measurement buffer and the impedimetric biosensing response has not been studied. Potentiometric biosensors were able to successfully detect the primary antibody and the target antigen by decreasing the ionic strength of the phosphate buffered saline (PBS) measurement buffer from 1x PBS to 0.01x PBS. However, the secondary antibody used for the selective signal amplification was not reliably detected. Therefore, the sandwich approach is not viable for potentiometric sensing at biologically relevant ionic strengths, due to the Debye screening effect. Alternatively, decreasing the ionic strength of the measurement buffer allowed for the successful translation of the sandwich approach to impedimetric biosensors. Impedimetric biosensing literature typically attributes a measured increase in the charge transfer resistance to an increase in the thickness of the immobilized biolayer. However, this work highlights the influence that both the charge and thickness of the biolayer have on the transport of the redox couple. Decreasing the ionic strength of the measurement buffer lowers the molecular charge screening effect. This permits the transport of a positively charged redox probe through a negatively charged immobilized biolayer via migration and diffusion. The results demonstrate that the use of a buffer at a lower, yet biologically relevant ionic strength allows for the successful translation of the sandwich approach to impedimetric biosensors.

Influence of the liquid ionic strength on the resonance frequency and shell parameters of lipid-coated microbubbles.

The correct measurement of the resonance frequency and shell properties of coated microbubbles (MBs) is essential in understanding and optimizing their response to ultrasound (US) exposure parameters. In diagnostic and therapeutic ultrasound, MBs are typically surrounded by blood; however, the influence of the medium charges on the MB resonance frequency has not been systematically studied using controlled measurements. This study aims to measure the medium charge interactions on MB behavior by measuring the frequency-dependent attenuation of the same size MBs in mediums with different charge densities. In-house lipid-coated MBs with C3F8 gas core were formulated. The MBs were isolated to a mean size of 2.35 µm using differential centrifugation. MBs were diluted to ≈8×105 MBs/mL in distilled water (DW), Phosphate-Buffered Saline solution (PBS1x) and PBS10x. The frequency-dependent attenuation of the MBs solutions was measured using an aligned pair of PVDF transducers with a center frequency of 10MHz and 100% bandwidth in the linear oscillation regime (7 kPa pressure amplitude). The MB shell properties were estimated by fitting the linear equation to experiments. Using a pendant drop tension meter, the surface tension at the equilibrium of ≈6 mm diameter size drops of the same MB shell was measured inside DW, PBS1x and PBS10x. The surface tension at the C3F8/solution interface was estimated by fitting the Young-Laplace equation from the recorded images. The frequency of the peak attenuation at different salinity levels was 13, 7.5 and 6.25 MHz in DW, PBS1x and PBS-10x, respectively. The attenuation peak increased by ≈140% with increasing ion density. MBs' estimated shell elasticity decreased by 64% between DW and PBS-1x and 36% between PBS-1x and PBS-10x. The drop surface tension reduced by 10.5% between DW and PBS-1x and by 5% between PBS-1x and PBS-10x, respectively. Reduction in the shell stiffness is consistent with the drop surface tension measurements. The shell viscosity was reduced by ≈40% between DW and PBS-1x and 42% between PBS-1x and PBS-10x. The reduction in the fitted stiffness and viscosity is possibly due to the formation of a densely charged layer around the shell, further reducing the effective surface tension on the MBs. The changes in the resonance frequency and estimated shell parameters were significant and may potentially help to better understand and explain bubble behavior in applications.