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Experimental Eye Research Aug 2023The single-cell RNA-sequencing (scRNA-seq) technique is used to explore the biological characteristics of tissues under pathological and physiological conditions that...
The single-cell RNA-sequencing (scRNA-seq) technique is used to explore the biological characteristics of tissues under pathological and physiological conditions that include certain chronic eye diseases. Harvesting of single-cell suspensions is one challenge inherent to scRNA-seq procedures. This study aimed to use an optimized method to digest a whole mouse cornea to harvest single-cell suspensions. We utilized five different mouse cornea digestion methods to obtain single-cell suspensions: (1) 5 dissected mouse corneas were cut into pieces (∼0.5 mm) and digested in trypsin for 10 min, and this digestion was repeated for 10 cycles; (2) 5 dissected mouse corneas were cut into pieces and incubated with 5 mg/ml collagenase A at 37 °C for 1h and then further digested in trypsin at 37 °C for 10 min; (3) used the same approach as that used in method 2, but the second digestion step was performed in TrypLE for 20 min; (4) used the same approach as that used in method 2, but the concentration of collagenase A was 2 mg/ml and the incubation time was 2h; (5) used the same approach as that used in method 3, but the corneas were incubated in 2 mg/ml collagenase A at 37 °C for 2h. Trypan blue staining was used to calculate the cell viability and agglomeration rate. The cell types and percentages were determined using immunofluorescence staining. RNA integrity number (RIN) was measured by Agilent 2100. Method 1 showed the lowest cell yield (0.375 × 10), epithelial cell percentage, and less than 70% cell viability, thus not a proper protocol. Method 2 showed the highest cell viability (over 90%), percentage of single-cell (89.53%), and high cell quantity (1.05 × 10). Method 3 had a significantly lower cell viability (55.30%). Cell agglomeration rates of method 4 and 5 reached up to 20% and 13%, and with lower cell viability (72.51%, 59.87%, respectively) and decreased epithelial cell rate compared to method 2 and 3. The results suggest that method 2 (5 mg/ml collagenase A and trypsin) is a preferred protocol for digesting mouse cornea to obtain single-cell suspension which achieves the criterion of single-cell RNA sequencing.
Topics: Mice; Animals; Trypsin; Suspensions; Collagenases; Cornea; RNA
PubMed: 37336469
DOI: 10.1016/j.exer.2023.109544 -
Chemosphere Nov 2020Intensive application of biochar requires better understanding of their environmental behaviors such as stability, fate, and mobility. The release of bulk biochar into...
Intensive application of biochar requires better understanding of their environmental behaviors such as stability, fate, and mobility. The release of bulk biochar into biochar nanoparticles (NPs) may bring risks because of their potential flowing into downstream water bodies with nutrients/containments attached. Low-temperature pyrolyzed biochars, namely fruit tree branch biochar of 350/450/550 °C (FB350, FB450 and FB550), corn straw biochar of 350 °C (CB350) and peanut straw biochar of 350 °C (PB350), were produced, and their NPs were extracted. The yield, elemental composition, mineral composition, surface functional groups and zeta potential of biochar NPs were characterized. Subsequently their suspension stability was evaluated in NaCl and CaCl solutions by dynamic light scattering technique. The Hamaker constants and particle interaction energy of the biochar NPs were calculated by adopting Derjaguin-Landau-Verwey-Overbeek theory. For biochar NPs of same feedstock, the stability of FB350/450/550-NPs could be predicted well by their zeta potential values. The types of their surface functional groups were the same while their adsorption intensity differed. The scenarios for biochar NPs of different feedstock sources were different, that is, inconsistent variation was observed between their zeta potential and suspension stability, which were rooted in the variable type and quantity of surface functional groups. In conclusion, feedstock was the most significant factor that influenced the suspension stability of biochar NPs, followed by the pyrolysis temperature and solution chemistry, which were highly dependent on surface potential. The findings provide references for the environmental risk evaluation of biochar NPs and reasonable application of biochar in field.
Topics: Adsorption; Charcoal; Dynamic Light Scattering; Nanoparticles; Pyrolysis; Solutions; Surface Properties; Suspensions; Temperature
PubMed: 32650172
DOI: 10.1016/j.chemosphere.2020.127510 -
Soft Matter Mar 2022A suspension of nanoparticles with very low volume fraction is found to assemble into a macroscopic cellular phase that is composed of particle-rich walls and...
A suspension of nanoparticles with very low volume fraction is found to assemble into a macroscopic cellular phase that is composed of particle-rich walls and particle-free voids under the collective influence of AC and DC voltages. Systematic study of this phase transition shows that it was the result of electrophoretic assembly into a two-dimensional configuration followed by spinodal decomposition into particle-rich walls and particle-poor cells mediated principally by electrohydrodynamic flow. This mechanistic understanding reveals two characteristics needed for a cellular phase to form, namely (1) a system that is considered two dimensional and (2) short-range attractive, long-range repulsive interparticle interactions. In addition to determining the mechanism underpinning the formation of the cellular phase, this work presents a method to reversibly assemble microscale continuous structures out of nanoscale particles in a manner that may enable the creation of materials that impact diverse fields including energy storage and filtration.
Topics: Electricity; Electrophoresis; Nanoparticles; Phase Transition; Suspensions
PubMed: 35080230
DOI: 10.1039/d1sm01650d -
The AAPS Journal Mar 2021The development of long-acting injectable (LAI) suspension products has increased in recent years. A better understanding of the relationship between the physicochemical...
The development of long-acting injectable (LAI) suspension products has increased in recent years. A better understanding of the relationship between the physicochemical properties of these products and their in vitro as well as in vivo performance is expected to further facilitate their development and regulatory review. Using Depo-SubQ Provera 104® as the reference listed drug (RLD), four qualitatively and quantitatively (Q1/Q2) equivalent LAI suspensions with different formulation properties were prepared. Two recrystallization methods (solvent evaporation and antisolvent) were utilized to obtain active pharmaceutical ingredient (API) with different properties and solid-state characterization was performed. In addition, two different sources of the major excipient were used to prepare the Q1/Q2 equivalent suspensions. Physiochemical characterization and in vitro release testing of the prepared Q1/Q2 equivalent suspension formulations and the RLD were conducted. In vitro drug release was dependent not only on the particle size, the morphology, and the crystallinity of the API but also on the residual solvent in the API. The excipient source also affected the drug release rates.
Topics: Chemistry, Pharmaceutical; Crystallization; Delayed-Action Preparations; Drug Compounding; Drug Liberation; Excipients; Injections, Intramuscular; Injections, Subcutaneous; Medroxyprogesterone Acetate; Particle Size; Solubility; Suspensions
PubMed: 33709196
DOI: 10.1208/s12248-021-00566-0 -
Zhongguo Zhong Yao Za Zhi = Zhongguo... Oct 2018Nanosuspension (also called nanocrystal suspension or nanocrystal) could significantly enhance the saturated solubility and dissolution of insoluble drugs, and improve... (Review)
Review
Nanosuspension (also called nanocrystal suspension or nanocrystal) could significantly enhance the saturated solubility and dissolution of insoluble drugs, and improve their bioavailability by reducing particle size and increasing the specific surface, which could then solve the delivery problems of the poorly soluble active ingredients and effective parts of Chinese materia medica (CMM). Based on the brief summaries of nanosuspension preparation methods, this paper would mainly review the and behaviors of poorly soluble CMM nanosuspension, discuss and analyze its problems, so as to provide reference and thinking for the further study of nanosuspension drug delivery system of poorly soluble CMM and promote the development and perfection of nanosuspension technology in CMM.
Topics: Biological Availability; Drug Delivery Systems; Materia Medica; Medicine, Chinese Traditional; Nanoparticles; Particle Size; Solubility; Suspensions
PubMed: 30453705
DOI: 10.19540/j.cnki.cjcmm.20180726.017 -
Journal of Colloid and Interface Science Dec 2022Capillary suspensions feature networks of particles connected by liquid bridges, which are obtained by adding a small amount of a second immiscible liquid to a...
HYPOTHESIS
Capillary suspensions feature networks of particles connected by liquid bridges, which are obtained by adding a small amount of a second immiscible liquid to a suspension. It is possible to link the network formation as well as the rheological behaviour of capillary suspensions to the intermolecular interactions of their constituents.
EXPERIMENTS AND SIMULATIONS
Through a combination of experimental and numerical methods, we present a novel approach, based on Hansen solubility parameters computed from Molecular Dynamics (MD) simulations, to rationalize and predict the rheological behaviour of capillary suspensions. We investigated the formation of capillary suspensions for various combinations of bulk and secondary liquids mixed with hydrophilic silica particles. The predictions were confirmed experimentally by rheological analysis, interfacial tension measurements and microscopy (CLSM) imaging.
FINDINGS
Numerical and experimental results show that the Hansen solubility parameters theory allows to predict the formation of capillary suspensions, whose strength exponentially decays with decreasing intermolecular interactions between the secondary liquids and the dispersed particles. High immiscibility between the bulk and secondary liquid strengthens the gel up to a critical immiscibility point, above which the strength of the gel remains mostly affected by the affinity between the secondary liquids and the dispersed particles. Furthermore, we find that hydrogen-bonding and polar interactions control the formation of capillary suspensions. This simple approach can guide the selection of adequate solvents and immiscible secondary liquids, allowing an easy formulation of new particulate-based gels.
Topics: Gels; Hydrogen; Silicon Dioxide; Solvents; Suspensions
PubMed: 35863200
DOI: 10.1016/j.jcis.2022.07.067 -
Molecules (Basel, Switzerland) May 2022Controllably accumulating and delivering nanoparticles (NPs) into specific locations are a central theme of nano-engineering and important for targeted therapy or...
Controllably accumulating and delivering nanoparticles (NPs) into specific locations are a central theme of nano-engineering and important for targeted therapy or bacteria removal. Here we present a technique allowing bidirectional accumulation, directional delivery and release of nanoparticles through two 980-nm-wavelength counter-propagating evanescent waves in an optical nanofiber (NF). Using 713-nm-diameter polystyrene NPs suspension and an 890-nm-diameter NF as an example, we experimentally and theoretically demonstrate that the NPs delivered along the NF surface in opposite directions are accumulated into the region where the scattering loss of the NPs is maximum, and about 90% of the incident optical field from both ends of the NF can be coupled into the region. Moreover, the accumulation region can be controlled by altering the incident optical power ratio of the two counter-propagating laser beams, while the accumulated NPs can be delivered and then released into the specific locations by turning off the two lasers.
Topics: Lasers; Nanofibers; Nanoparticles; Polystyrenes; Suspensions
PubMed: 35630790
DOI: 10.3390/molecules27103312 -
Journal of Pharmaceutical Sciences Jan 2023High concentration protein formulations for subcutaneous injection represent a substantial number of development projects in the pharmaceutical industry. Such...
High concentration protein formulations for subcutaneous injection represent a substantial number of development projects in the pharmaceutical industry. Such concentrated aqueous protein solutions face some specific challenges such as increased viscosity and aggregation propensity. Protein powder suspensions in non-aqueous vehicles could be an alternative providing lower viscosity than the respective aqueous solution. The choice of potential suspension vehicles is limited as traditional non-aqueous liquids, such as oils, show an inherent high viscosity. We studied suspensions prepared by dispersing spray-dried protein powder in different vehicles including sesame oil and medium chain triglycerides, as well as fluorinated and semifluorinated alkanes. We found, that semifluorinated alkanes enable formulations with high concentrations up to 280 mg/ml monoclonal antibody with a low viscosity of less than 10 mPa·s and low injection forces. The glide force of suspensions containing 210 mg/ml protein was not affected by the particle size of the spray-dried powders with medians ranging from 1 to 14 µm. In contrast, suspensions prepared with cryo-milled powder showed markedly higher viscosities and were not injectable at the same concentration. Protein powder suspensions were syringeable using a 25G needle. Vial filling using a peristaltic pump was possible and lead to a uniform filling. Sedimentation of the suspension was slow and does not lead to challenges upon vial filling during manufacturing or transfer of the suspension into syringes. Thus, we could show that dispersions of spray-dried protein powders in non-aqueous vehicles, such as semifluorinated alkanes, are a promising alternative to aqueous protein solutions at high concentrations.
Topics: Powders; Suspensions; Particle Size; Viscosity; Excipients; Alkanes
PubMed: 35779665
DOI: 10.1016/j.xphs.2022.06.016 -
AAPS PharmSciTech Jan 2018This study examined the use of focused beam reflectance measurement (FBRM) for qualitative and quantitative analysis of pharmaceutical suspensions with particular...
This study examined the use of focused beam reflectance measurement (FBRM) for qualitative and quantitative analysis of pharmaceutical suspensions with particular application to toxicology supply preparations for use in preclinical studies. Aqueous suspensions of ibuprofen were used as prototype formulations. Initial experiments were conducted to examine the effects of operational conditions including FBRM probe angle, probe location, and mixing (method and rate of mixing) on the FBRM analysis. Once experimental conditions were optimized, the homogeneity and sedimentation-redispersion of particles in the suspensions were assessed. Ibuprofen suspension under continuous agitation was monitored using FBRM for 60 h to study particle size change over time. Another study was performed to determine if particle count rates obtained by FBRM could be correlated to suspension concentration. The location and the angle of the FBRM probe relative to the beaker contents, and the rate and the method of mixing the suspension were found to be sensitive parameters during FBRM analysis. FBRM was able to monitor the process of particle sedimentation in the suspension. The attrition of ibuprofen particles was detectable by FBRM during prolonged stirring with an increase in the number of smaller particles and decrease in the number of larger particles. A strong correlation was observed between particle count rate by FBRM and ibuprofen concentration in the suspension. Also, change in content uniformity in the suspension at different locations of the beaker was represented by FBRM particle count. Overall, FBRM has potential to be a useful tool for qualitative and quantitative analysis of pharmaceutical suspensions.
Topics: Feasibility Studies; Ibuprofen; Particle Size; Suspensions; Technology, Pharmaceutical
PubMed: 28639177
DOI: 10.1208/s12249-017-0819-9 -
Water Research Nov 2022Interplays between microalgae and clay minerals enhance biologically mediated flocculation, thereby affecting the sedimentation and transportation of suspended...
Interplays between microalgae and clay minerals enhance biologically mediated flocculation, thereby affecting the sedimentation and transportation of suspended particulate matter (SPM) in water and benthic environments. This interaction forms larger flocs with a higher settling velocity and enhances SPM sinking. The aim of this study was to investigate the flocculation kinetics of microalgae and clay in suspension and to elucidate the mechanisms associated with such interactions. Standard jar test experiments were conducted using various mixtures of kaolinite and microalgal samples from batch cultures (Chlorella vulgaris) to estimate biologically mediated flocculation kinetics. The organic matter (OM) composition secreted by the microalgae was characterized using a liquid chromatography - organic carbon detection system, and quantitative analysis of transparent exopolymer particles was conducted separately. A two-class flocculation kinetic model, based on the interaction between flocculi and flocs, was also adopted to quantitatively analyze the experimental data from flocculation. Results from the flocculation kinetic tests and OM analyses, in association with other data analyses (i.e., floc size distribution and flocculation kinetic model), showed that flocculation increased with OM concentration during the growth phase (10-20 d). However, on day 23 during the early stationary phase, flocculation kinetics started decreasing and substantially declined on day 30, even though the amount of OM (mainly biopolymers) continued to increase. Our results indicate that an adequate quantity of biopolymers produced by the microalgal cells in the growth phase enhanced floc-to-floc attachment and hence flocculation kinetics. In contrast, an excessive quantity of biopolymers and humic substances in the stationary phase enhanced the formation of polymeric backbone structures and flocculation via scavenging particles but simultaneously increased steric stabilization with the production of a large number of fragmented particles.
Topics: Flocculation; Microalgae; Clay; Chlorella vulgaris; Suspensions; Kinetics
PubMed: 36323221
DOI: 10.1016/j.watres.2022.119300