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In patients with total hip replacements (THRs), wear products in the form of nanoparticles and ions are released, especially around implant failure. In this study, we use N2a cells, a neuroblastoma cell line, to evaluate the effects of different flow rates on neuronal toxicity amidst exposure to CoCrMo particles. We hypothesized that increasing flow rates would increase N2a cell viability and decrease N2a cell-degradation products (DPs) toxicity. We conducted four 24-hour experiments, each with four flow rate conditions, 0, 50, 100, and 200 μL/min, based on the physiological shear stress of the vessels in the human body, to evaluate cell viability, cell morphology, and cell-DPs interaction. Steps included microfluidic channel preparation, N2a cell culturing, CoCrMo particle acquisition, microfluidic system assembly, and dynamic flow neurotoxicity evaluation, which included video microscopy, AlamarBlue, live/dead imaging, DAPI, and ROS assay. The results suggest that fewer neurotoxic reactions and greater viability at higher flow rates supported our hypothesis, although the full range of viable flow rates is yet to be studied. While cell-particle interaction is complex and dynamic, flow rate did modulate toxicity, viability, morphology, and growth environment. The microfluidic system should continue to be developed to study toxicology aspects of implants by simulating in vivo conditions more accurately. [Display omitted] • Neuronal toxicity risks associated with CoCrMo particles total hip replacement (THR) are studied utilizing microfluidics. • This study highlights influence of fluid dynamics on cell-particle interactions, the impact on toxicity pathways. • This study is the first demonstration of microfluidic technology to mimic systemic toxicity from implants. • The findings indicate that increasing flow rates result in a reduction of cell aggregation and particle sedimentation.. • This in turn leads to a decrease in neurotoxicity and an improvement in cell viability. • Possibility of using the developed microfluidic device to other applications, such as cardio-vascular diseases. [ABSTRACT FROM AUTHOR]

Artificial intelligence (AI)-guided methods are transforming the speed and scale with which image segmentation and classification tasks can be managed in cell biology. Deep learning (DL)-guided tools to segment and classify a variety of cells and subcellular structures are being rapidly developed, opening the need for standards and repositories. Despite DL-guided advances in still-image analysis, tracking objects in microscopy movies remains an area of open development owing to spatial and temporal discontinuities. DL methods offer new opportunities to significantly expand genotype–phenotype maps, genetic variant analysis, and drug development and discovery. The growth of artificial intelligence (AI) has led to an increase in the adoption of computer vision and deep learning (DL) techniques for the evaluation of microscopy images and movies. This adoption has not only addressed hurdles in quantitative analysis of dynamic cell biological processes but has also started to support advances in drug development, precision medicine, and genome–phenome mapping. We survey existing AI-based techniques and tools, as well as open-source datasets, with a specific focus on the computational tasks of segmentation, classification, and tracking of cellular and subcellular structures and dynamics. We summarise long-standing challenges in microscopy video analysis from a computational perspective and review emerging research frontiers and innovative applications for DL-guided automation in cell dynamics research. The growth of artificial intelligence (AI) has led to an increase in the adoption of computer vision and deep learning (DL) techniques for the evaluation of microscopy images and movies. This adoption has not only addressed hurdles in quantitative analysis of dynamic cell biological processes but has also started to support advances in drug development, precision medicine, and genome–phenome mapping. We survey existing AI-based techniques and tools, as well as open-source datasets, with a specific focus on the computational tasks of segmentation, classification, and tracking of cellular and subcellular structures and dynamics. We summarise long-standing challenges in microscopy video analysis from the computational perspective and review emerging research frontiers and innovative applications for DL-guided automation in cell dynamics research. [ABSTRACT FROM AUTHOR]

Unlike the mechanism of ballistospore discharge, which was not solved until the 1980s, the operation of asci as pressurized squirt guns is relatively straightforward and was understood in the nineteenth century. Since then, mycologists have sought to understand how structural adaptations to asci have allowed the ascomycetes to expel spores of different shapes and sizes over distances ranging from a few millimeters to tens of centimeters. These modifications include the use of valves at the tips of asci that maintain ascus pressure and expel spores at the highest speeds, and gelatinous appendages that connect spores after release and create larger projectiles with greater momentum than single spores. Clever experiments in the twentieth century coupled with meticulous microscopic studies led investigators to understand how asci with complicated apical structures worked and mathematical models produced estimates of launch speeds. With the recent application of high-speed video microscopy, these inferences about ascus function have been tested by imaging the motion of spores on a microsecond timescale. These experiments have established that ascospore discharge is the fastest fungal movement and is among the fastest movements in biology. Beginning with the history of the study of asci, this review article explains how asci are pressurized, how spores are released, and how far spores travel after their release. We also consider the efficiency of ascospore discharge relative to the mechanism of ballistospore discharge and examine the way that the squirt gun mechanism has limited the morphological diversity of ascomycete fruit bodies. • Ascospore discharge is the fastest fungal movement. • Early ascus research inspired the most ingenious experiments in mycological history. • Ascospore and ballistospore discharge share the same underlying chemistry. [ABSTRACT FROM AUTHOR]

Mechanical stress patterns emerging from collective cell behavior have been shown to play critical roles in morphogenesis, tissue repair, and cancer metastasis. In our previous work, we constrained valvular interstitial cell (VIC) monolayers on circular protein islands to study emergent behavior in a controlled manner and demonstrated that the general patterns of cell alignment, size, and apoptosis correlate with predicted mechanical stress fields if radially increasing stiffness or contractility are used in the computational models. However, these radially symmetric models did not predict the existence of local regions of dense aligned cells observed in seemingly random locations of individual aggregates. The goal of this study is to determine how the heterogeneities in cell behavior emerge over time and diverge from the predicted collective cell behavior. Cell-cell interactions in circular multicellular aggregates of VICs were studied with time-lapse imaging ranging from hours to days, and migration, proliferation, and traction stresses were measured. Our results indicate that elongated cells create strong local alignment within preconfluent cell populations on the microcontact printed protein islands. These cells influence the alignment of additional cells to create dense, locally aligned bands of cells which disrupt the predicted global behavior. Cells are highly elongated at the endpoints of the bands yet have decreased spread area in the middle and reduced mobility. Although traction stresses at the endpoints of bands are enhanced, even to the point of detaching aggregates from the culture surface, the cells in dense bands exhibit reduced proliferation, less nuclear YAP, and increased apoptotic rates indicating a low stress environment. These findings suggest that strong local cell-cell interactions between primary fibroblastic cells can disrupt the global collective cellular behavior leading to substantial heterogeneity of cell behaviors in constrained monolayers. This local emergent behavior within aggregated fibroblasts may play an important role in development and disease of connective tissues. Mechanical stress patterns emerging from collective cell behavior play critical roles in morphogenesis, tissue repair, and cancer metastasis. Much has been learned of these collective behaviors by utilizing microcontact printing to constrain cell monolayers (aggregates) into specific shapes. Here we utilize these tools along with long-term video microscopy tracking of individual aggregates to determine how heterogeneous collective behaviors unique to primary fibroblastic cells emerge over time and diverge from computed stress fields. We find that dense multicellular bands form from local collective behavior and disrupt the global collective behavior resulting in heterogeneous patterns of migration, traction stresses, proliferation, and apoptosis. This local emergent behavior within aggregated fibroblasts may play an important role in development and disease of connective tissues. [Display omitted] [ABSTRACT FROM AUTHOR]

Ultrasound (US)–targeted microbubble (MB) cavitation (UTMC)–mediated therapies have been found to restore perfusion and enhance drug/gene delivery. Because of the potentially longer circulation time and relative ease of storage and reconstitution of polymer-shelled MBs compared with lipid MBs, we investigated the dynamic behavior of polymer microbubbles and their therapeutic potential for sonoreperfusion (SRP) therapy. The fate of polymer MBs during a single long tone-burst exposure (1 MHz, 5 ms) at various acoustic pressures and MB concentrations was recorded via high-speed microscopy and passive cavitation detection (PCD). SRP efficacy of the polymer MBs was investigated in an in vitro flow system and compared with that of lipid MBs. Microscopy videos indicated that polymer MBs formed gas-filled clusters that continued to oscillate, fragment and form new gas-filled clusters during the single US burst. PCD confirmed continued acoustic activity throughout the 5-ms US excitation. SRP efficacy with polymer MBs increased with pulse duration and acoustic pressure similarly to that with lipid MBs but no significant differences were found between polymer and lipid MBs. These data suggest that persistent cavitation activity from polymer MBs during long tone-burst US excitation confers excellent reperfusion efficacy. [ABSTRACT FROM AUTHOR]

Determination of subvisible particle (SVP) content in biopharmaceuticals is a prerequisite to ensure the quality of liquid biopharmaceutical products. Here, we present a comparison of the recently introduced holographic video microscopy (total holographic characterization, THC) with two orthogonal and well-established analytical technologies: micro flow imaging (MFI) and resonant mass measurement (RMM). The capabilities of the THC were investigated under conditions commonly applied in drug product development. Three different antibody products were used at different concentrations and formulations to cover a wide range of realistic use-cases. The comparison was particularly focused on protein aggregates to investigate the applicability of THC to this critical class of particles in drug product development. Protein concentrations up to 100 mg/ml were investigated covering a broad range of viscosity and refractive indices, both important parameters in particle detection. The comparison reveals that THC is highly sensitive to detect protein aggregates in a size range from 0.5 µm to 10 µm. THC shows a significant superiority to FI and RMM in detecting heterogenous protein aggregates which often appear as transparent and porous particles. Additionally, THC needs very small sample amount of about 30 µl and short measurement times, making it applicable for early development stages and high-throughput approaches. These results show that THC is a valuable supplement to the existing particle characterization method portfolio in drug product development. [ABSTRACT FROM AUTHOR]

Mitochondria are dynamic organelles essential for cellular homeostasis, undergoing continuous fission and fusion processes that regulate their morphology, distribution, and function. Disruptions in these dynamics are linked to numerous diseases, including neurodegenerative disorders and cancer. Understanding these processes is vital for developing therapeutic strategies aimed at mitigating mitochondrial dysfunction. This review provides an overview of recent perspectives on mitochondrial dynamics, focusing on the need for live video microscopy imaging in order to fully understand mitochondrial phenotypes and pathology. Advanced imaging tools, such as machine learning-based segmentation and label-free microscopy approaches, have the potential to transform our ability to study mitochondrial dynamics in live cells. • Mitochondrial fission and fusion processes are tightly regulated and interconnected. • How fission/fusion dynamics and retrograde/anterograde movement are balanced cannot be captured with single timepoint images. • Advanced imaging tools that capture spatiotemporal dynamics are thus necessary for understanding mitochondrial behavior. [ABSTRACT FROM AUTHOR]

• Ionic liquid enhances evaporation of a water droplet by assembling at the water–air interface. • Ionic liquid reduces interfacial tension enhancing water diffusion from liquid to gas phase. • Low solvation energy assembles ionic liquids at interface to control water evaporation kinetics. Evaporation of a water droplet is well understood from the view of molecular diffusion. Even though there are reports that discuss the effects of inorganic salts, the evaporation of a water droplet with added organic salts, such as ionic liquids, is rare. In the present study, ionic liquids controlled simultaneous spreading and evaporation of a water droplet have been studied using video microscopy techniques. In the presence of these molecules, the sessile drop starts to evaporate in a constant contact radius (CCR) mode, due to the pinning of the triple-phase contact line to a hydrophobic substrate. Later, it shifts to constant contact angle (CCA) mode for a short while, which finally enters a contact angle recovery mode after reaching to a minimal. The presence of these molecules in water ultimately enhances the evaporation rate by assembling themselves at the water–air interface. Such assembly reduces the interfacial tension enhancing the probability of escaping the water molecule from liquid to gas phase. The variation of volume with time has been fitted using a model equation which indicates the increased value of diffusion coefficient. Additionally, the experiments with varying relative humidity show that the contact angle recovery is prominent in a humid environment. The molecular dynamic simulation has been performed to finally calculate the solvation free energy of the system, which ensures the ionic liquids to assemble at the interface. This assembly plays a vital role in evaporation kinetics. [ABSTRACT FROM AUTHOR]

The inherently dynamic and anisotropic microenvironment of cells imposes not only global and slow physical stimulations on cells but also acute and local perturbations. However, cell mechanical responses to transient subcellular physical signals remain unclear. In this study, acoustically activated targeted microbubbles were used to exert mechanical perturbations to single cells. The cellular contractile force was sensed by elastic micropillar arrays, while the pillar deformations were imaged using brightfield high-speed video microscopy at a frame rate of 1k frames per second for the first 10s and then confocal fluorescence microscopy. Cell mechanical responses are accompanied by cell membrane integrity changes. Both processes are determined by the perturbation strength generated by microbubble volumetric oscillations. The instantaneous cellular traction force relaxation exhibits two distinct patterns, correlated with two cell fates (survival or permanent damage). The mathematical modeling unveils that force-induced actomyosin disassembly leads to gradual traction force relaxation in the first few seconds. The perturbation may also influence the far end subcellular regions from the microbubbles and may propagate into connected cells with attenuations and delays. This study carefully characterizes the cell mechanical responses to local perturbations induced by ultrasound and microbubbles, advancing our understanding of the fundamentals of cell mechano-sensing, -responsiveness, and -transduction. Subcellular physical perturbations commonly exist but haven't been fully explored yet. The subcellular perturbation generated by ultrasound and targeted microbubbles covers a wide range of strength, from mild, intermediate to intense, providing a broad biomedical relevance. With µm2 spatial sensing ability and up to 1ms temporal resolution, we present spatiotemporal details of the instantaneous cellular contractile force changes followed by attenuated and delayed global responses. The correlation between the cell mechanical responses and cell fates highlights the important role of the instantaneous mechanical responses in the entire cellular reactive processes. Supported by mathematical modeling, our work provides new insights into the dynamics and mechanisms of cell mechanics. [Display omitted] [ABSTRACT FROM AUTHOR]

Today, sodium glucose co-transporter 2 (SGLT2) inhibitors are more than diabetes drugs. They are also indicated in chronic heart failure (HF) treatment in both diabetic and non-diabetic patients, independently of the ejection fraction. Multiple mechanisms have been suggested behind the cardioprotective effects of SGLT2 inhibitors. However, the underlying mechanisms still remain largely unexplored. Here, we used a zebrafish embryo model to search for new potential players whereby SGLT2 inhibitors attenuate HF. HF in zebrafish embryos was caused exposing them to chemically induced hypoxia. As a SGLT2 inhibitor, we used empagliflozin. Its effect on hypoxia-induced HF of the embryos was evaluated using video microscopy and calculation of fractional shortening (FS) of embryos´ hearts. RT-qPCR of brain natriuretic peptide (bnp) expression was also used to examine empagliflozin´s effect on HF. Transcriptome analysis of total RNA of the embryos was performed to search for new potential mechanisms contributing to the beneficial effect of empagliflozin on HF. Empagliflozin significantly attenuated hypoxia-induced HF of zebrafish embryos as shown with improved FS of the hearts and decreased bnp expression. Transcriptome analysis revealed that the improvement of HF in response to empagliflozin was accompanied with decreased matrix metalloproteinase 13a (mmp13a) expression. Treatment of hypoxia-induced embryos with MMP13 inhibitor ameliorated the impaired heart function accordingly to the effect of empagliflozin. MMP13 inhibitor was not toxic to the embryos. Our study shows that empagliflozin´s favorable effect on attenuating HF is mediated via MMP13. MMP13 provides a novel option when developing new therapeutics for HF treatment. • Empagliflozin attenuates hypoxia-induced heart failure (HF) of zebrafish embryos. • Empagliflozin´s beneficial effect on HF of zebrafish embryos is mediated via MMP13. • MMP13 provides a novel option in developing new therapeutics for HF treatment. [ABSTRACT FROM AUTHOR]

Plasmin is a broad-spectrum protease and therefore needs to be tightly regulated. Active plasmin is formed from plasminogen, which is found in high concentrations in the blood and is converted by the plasminogen activators. In the circulation, high levels of a2-antiplasmin rapidly and efficiently inhibit plasmin activity. Certain myeloid immune cells have been shown to bind plasmin and plasminogen on their cell surface via proteins that bind to the plasmin(ogen) kringle domains. Our earlier work showed that T cells can activate plasmin but that they do not themselves express plasminogen. Here, we demonstrate that T cells express several known plasminogen receptors and that they bind plasminogen on their cell surface. We show T cell-bound plasminogen was converted to plasmin by plasminogen activators upon T cell activation. To examine functional consequences of plasmin generation by activated T cells, we investigated its effect on the chemokine, C-C motif chemokine ligand 21 (CCL21). Video microscopy and Western blotting confirmed that plasmin bound by human T cells cleaves CCL21 and increases the chemotactic response of monocyte-derived dendritic cells toward higher CCL21 concentrations along the concentration gradient by increasing their directional migration and track straightness. These results demonstrate how migrating T cells and potentially other activated immune cells may co-opt a powerful proteolytic system from the plasma toward immune processes in the peripheral tissues, where a2-antiplasmin is more likely to be absent. We propose that plasminogen bound to migrating immune cells may strongly modulate chemokine responses in peripheral tissues. [ABSTRACT FROM AUTHOR]

• Intracellular protist parasites have similar basal Ca2+ levels than their host cells. • Ca2+ signaling faciitates host cell invasion, adaptation to intracellular life, and organell regulation. • Trypanosomatids possess well defined IP 3 receptors. • There is no genomic evidence for typical IP 3 or ryanodine receptors in apicomplexan parasites. • GECIs and video microscopy are being used to follow signaling in intracellular parasites. Calcium ion (Ca2+) signaling is one of the most frequently employed mechanisms of signal transduction by eukaryotic cells, and starts with either Ca2+ release from intracellular stores or Ca2+ entry through the plasma membrane. In intracellular protist parasites Ca2+ signaling initiates a sequence of events that may facilitate their invasion of host cells, respond to environmental changes within the host, or regulate the function of their intracellular organelles. In this review we examine recent findings in Ca2+ signaling in two groups of intracellular protist parasites that have been studied in more detail, the apicomplexan and the trypanosomatid parasites. [ABSTRACT FROM AUTHOR]

To ensure the stability of coal roadway props, the roadside support filling material must not only provide sufficient strength but also exhibit significant compression deformation capacity. Traditional concrete, primarily used in the construction of roads, bridges, and housing, emphasizes strength design with little consideration for deformation performance, rendering it unsuitable for the ultra-high pressure plastic deformation demands of roadside support filling materials in mining areas. This study prepared fiber-reinforced spherical aggregates through granulation molding of freshly mixed cement slurry with embedded fibers, and examined the influence of fiber content and size on the cylinder compressive strength and compression deformation of the spherical aggregates. The mechanical and compressive plastic deformation properties of fiber spherical aggregate concrete (FSAC) were investigated through compressive strength, flexural strength, and stress-strain curves. Additionally, video electron microscopy and SEM analysis were employed to explore the internal mechanisms by which fiber spherical aggregates (FSA) enhance the compressive plastic deformation properties of concrete. The test results indicate that although the cylinder compressive strength of FSA is marginally lower than that of natural aggregate (NA), it exhibits exceptional plastic deformation capacity. With the increment in fiber content or size, the compressive strength of FSAC remains relatively unchanged compared to NAC. However, the flexural strength is markedly enhanced when the fiber content reaches 1.5 % and the fiber length is 9 mm. Unlike the brittle failure observed in natural aggregate concrete (NAC) after crushing, FSAC specimens maintain good integrity post-compression failure. Additionally, FSAC demonstrates higher peak stress and stable strain near the peak stress, indicative of pronounced plastic deformation characteristics. The plasticity change in FSAC primarily relies on the exceptional compression plastic transformation properties of the FSA, which convert point stress at aggregate contacts into flexible surface contact, thereby dispersing stress and enhancing the overall compressive deformation capacity of the concrete. • This study prepared fiber-reinforced spherical aggregates through granulation molding of freshly mixed cement slurry with embedded fibers, and examined the influence of fiber content and size on the cylinder compressive strength and compression deformation of the spherical aggregates. The mechanical and compressive plastic deformation properties of fiber spherical aggregate concrete (FSAC) were investigated through compressive strength, flexural strength, and stress-strain curves. • With the increase in fiber content or length, the cylinder compressive strength of spherical aggregate exhibited an initial increase followed by a decrease. When the fiber length is 9 mm and the content is 1.5 %, the cylinder compressive strength of the spherical aggregate reaches 65.09 MPa, approximately 80 % of that of natural aggregate (NA). Concurrently, the fiber spherical aggregate (FSA) also demonstrates exceptional compression deformation characteristics. • The compressive strength of FSAC remains relatively unchanged compared to natural aggregate concrete (NAC). However, compared to NAC and spherical aggregate concrete without fibers, the flexural strength of FSAC is markedly enhanced. When the incorporated fiber length is 9 mm and the content is 1.5 %, the flexural strength of FSAC increases by 52.2 % compared to NAC. • Unlike the brittle failure of NAC upon crushing, FSAC exhibits more vertical cracks on the surface after compression failure, along with greater lateral expansion deformation. The crushed specimen maintains good integrity, characteristic of typical plastic failure. Compared to NAC, FSAC exhibits a higher peak stress and peak strain, along with a strain stability stage near the peak stress, indicative of clear plastic deformation characteristics. • When subjected to external loads, FSAC utilizes the excellent compression and plastic properties of FSA to convert the point stress, resulting from aggregate contact, into flexible surface contact. This dispersion of stress enhances the overall compression deformation capacity of the concrete. [ABSTRACT FROM AUTHOR]

Flagellum-mediated motility is essential to Pseudomonas aeruginosa (P. aeruginosa) virulence. Antibody against flagellin reduces motility and inhibits the spread of the bacteria from the infection site. The standard soft-agar assay to demonstrate anti-flagella motility inhibition requires long incubation times, is difficult to interpret, and requires large amounts of antibody. We have developed a time-lapse video microscopy method to analyze anti-flagellin P. aeruginosa motility inhibition that has several advantages over the soft agar assay. Antisera from mice immunized with flagellin type A or B were incubated with Green Fluorescent Protein (GFP)-expressing P. aeruginosa strain PAO1 (FlaB+) and GFP-expressing P. aeruginosa strain PAK (FlaA+). We analyzed the motion of the bacteria in video taken in ten second time intervals. An easily measurable decrease in bacterial locomotion was observed microscopically within minutes after the addition of small volumes of flagellin antiserum. From data analysis, we were able to quantify the efficacy of anti-flagellin antibodies in the test serum that decreased P. aeruginosa motility. This new video microscopy method to assess functional activity of anti-flagellin antibodies required less serum, less time, and had more robust and reproducible endpoints than the standard soft agar motility inhibition assay. [ABSTRACT FROM AUTHOR]

• Crystallization of bulk Bioglass 45S5 has been imaged from nano- to microscale. • Combining ex situ and in situ techniques allows to follow the entire crystallization process. • Combeite crystallizes with a sphere-like morphology visualized by nano-CT. • Combeite spheres present a grain-like internal structure. • Crystallinity of powder and bulk samples have been correlated with the images. One key issue influencing a broader application of Bioglass 45S5 in tissue engineering is its inherent crystallization tendency, severely limiting the mechanical strength of 3D porous scaffolds. Despite numerous studies, Bioglass 45S5 crystallization is not yet fully understood with regard to the mechanisms involved or morphology of the crystal phases forming. Here we show how two cutting-edge imaging techniques, state-of-the-art transmission electron microscopy (TEM) with image correction including energy dispersive X-ray spectroscopy and X-ray nano-computed tomography (nano-CT), allowed us to visualize changes in microstructure from near-nucleation to almost full crystallization in bulk Bioglass 45S5. At early times of heat treatment at 660 °C the formation of phase-separated nano-droplets within the glassy matrix was observed. Later, besides surface crystallization, bulk crystallization of combeite spheres was predominant. The formation of the first combeite spheres, their coarsening with time and finally their merging at near full crystallization were recorded by in situ high-temperature optical microscopy videos. The 3D nature of these spheres was confirmed by nano-CT, while TEM showed that their internal structure was composed of sub-micron grains. X-ray diffraction analysis at early time points showed a much higher crystalline fraction in bulk samples compared to powder samples, highlighting the influence of processing and sample morphology. These results show the importance of using complementary techniques for gaining insight into the crystallization process in the volume. In addition, we show that TEM and nano-CT are suitable characterization techniques to visualize the crystallization even in fast crystallizing systems, such as bioactive glasses. [ABSTRACT FROM AUTHOR]

A significant proportion of urinary tract infection (UTI) patients experience recurrent episodes, due to deep tissue infection and treatment-resistant bacterial reservoirs. Direct bladder instillation of antibiotics has proved disappointing in treating UTI, likely due to the failure of infused antibiotics to penetrate the bladder epithelium and accumulate to high enough levels to kill intracellular bacteria. This work investigates the use of nitrofurantoin loaded poly(lactic- co -glycolic acid) (PLGA) particles to improve delivery to intracellular targets for the treatment of chronic UTI. Using electrohydrodynamic atomisation, we produced particles with an average diameter of 2.8 μm. In broth culture experiments, the biodegradable particles were effective against a number of UTI-relevant bacterial strains. Dye-loaded particles demonstrated that intracellular delivery was achieved in all cells in 2D cultures of a human bladder epithelial progenitor cell line in a dose-dependent manner, achieving far higher efficiency and concentration than equivalent quantities of free drug. Time-lapse video microscopy confirmed that delivery occurred within 30 min of administration, to 100% of cells. Moreover, the particles were able to deliver the drug to cells through multiple layers of a 3D human bladder organoid model causing minimal cell toxicity, displaying superior killing of bacterial reservoirs harboured within bladder cells compared with unencapsulated drug. The particles were also able to kill bacterial biofilms more effectively than the free drug. These results illustrate the potential for using antibiotic-loaded microparticles to effectively treat chronic UTIs. Such a delivery method could be extrapolated to other clinical indications where robust intracellular delivery is required, such as oncology and gene therapy. Unlabelled Image [ABSTRACT FROM AUTHOR]

We demonstrate the use of holographic video microscopy to detect individual subvisible particles dispersed in biopharmaceutical formulations and to differentiate them based on material characteristics measured from their holograms. The result of holographic analysis is a precise and accurate measurement of the concentrations and size distributions of multiple classes of subvisible contaminants dispersed in the same product simultaneously. We demonstrate this analytical technique through measurements on model systems consisting of human IgG aggregates in the presence of common contaminants such as silicone oil emulsion droplets and fatty acids. Holographic video microscopy also clearly identifies metal particles and air bubbles. Being able to differentiate and characterize the individual components of such heterogeneous dispersions provides a basis for tracking other factors that influence the stability of protein formulations including handling and degradation of surfactant and other excipients. [ABSTRACT FROM AUTHOR]

• Structural dissimilarity analysis provides structural dynamics maps from videos. • Structural dissimilarity analysis is segmentation-free analysis method but provides comparable results to segmentation analysis. • Structural dissimilarity analysis is simple and computationally efficient. In-situ liquid cell transmission electron microscopy (LCTEM) with electrical biasing capabilities has emerged as an invaluable tool for directly imaging electrode processes with high temporal and spatial resolution. However, accurately quantifying structural changes that occur on the electrode and subsequently correlating them to the applied stimulus remains challenging. Here, we present structural dissimilarity (DSSIM) analysis as segmentation-free video processing algorithm for locally detecting and quantifying structural change occurring in LCTEM videos. In this study, DSSIM analysis is applied to two in-situ LCTEM videos to demonstrate how to implement this algorithm and interpret the results. We show DSSIM analysis can be used as a visualization tool for qualitative data analysis by highlighting structural changes which are easily missed when viewing the raw data. Furthermore, we demonstrate how DSSIM analysis can serve as a quantitative metric and efficiently convert 3-dimensional microscopy videos to 1-dimenional plots which makes it easy to interpret and compare events occurring at different timepoints in a video. In the analyses presented here, DSSIM is used to directly correlate the magnitude and temporal scale of structural change to the features of the applied electrical bias. ImageJ, Python, and MATLAB programs, including a user-friendly interface and accompanying documentation, are published alongside this manuscript to make DSSIM analysis easily accessible to the scientific community. [ABSTRACT FROM AUTHOR]

• CNN based framework for cell instance segmentation and tracking. • Incorporating temporal information as ConvGRUs into a fully convolutional network. • Embedding loss based on cosine similarities to represent individual instances. • Extensive evaluation on muscle fiber and celltracking microscopy images. • State-of-the-art performance on six datasets of the ISBI celltracking challenge. Differently to semantic segmentation, instance segmentation assigns unique labels to each individual instance of the same object class. In this work, we propose a novel recurrent fully convolutional network architecture for tracking such instance segmentations over time, which is highly relevant, e.g., in biomedical applications involving cell growth and migration. Our network architecture incorporates convolutional gated recurrent units (ConvGRU) into a stacked hourglass network to utilize temporal information, e.g., from microscopy videos. Moreover, we train our network with a novel embedding loss based on cosine similarities, such that the network predicts unique embeddings for every instance throughout videos, even in the presence of dynamic structural changes due to mitosis of cells. To create the final tracked instance segmentations, the pixel-wise embeddings are clustered among subsequent video frames by using the mean shift algorithm. After showing the performance of the instance segmentation on a static in-house dataset of muscle fibers from H&E-stained microscopy images, we also evaluate our proposed recurrent stacked hourglass network regarding instance segmentation and tracking performance on six datasets from the ISBI celltracking challenge, where it delivers state-of-the-art results. [ABSTRACT FROM AUTHOR]

• HIP W/O emulsion enhances the flotation of fine oxidized coal at low dosage. • Stable W 1 /O/W 2 double emulsion is formed when HIP emulsion is dispersed in water. • Emulsion droplets dispersed in water attach and spread at the oxidized coal surface. • H-bonds between Span 80 and coal surface expected to facilitate droplet attachment. Flotation recovery of fine oxidized coal is frequently inefficient when using conventional oily collectors. A large amount of fine coal is discharged as tailings, leading to resource waste and environmental pollution. To solve the problem of high oil consumption and low efficiency associated with traditional flotation, research efforts have been focused on the use of oil-in-water emulsion to reduce oil consumption. Few studies concerning water-in-oil emulsion, especially the high internal phase water-in-oil (HIP W/O) emulsion, have been conducted. Our microflotation results showed a significant oil dosage reduction when a HIP W/O emulsion was used as a collector for coal flotation. The state of the HIP W/O emulsion dispersion in water and the emulsion droplets attachment at the coal surface are vital issues associated with achieving efficient flotation. In this study, optical microscopy and high speed video microscopy were employed to study the dispersion behavior and microstructure of the HIP W/O emulsion in water. This study found that the HIP emulsion tends to be dispersed as a W 1 /O/W 2 double emulsion during mixing in water because of the migration of the surplus Span 80 in the oil phase to the O-W 2 interface. The presence of NaCl in W 1 is also essential for a stable W 1 /O/W 2 double emulsion, which is formed when equilibrium of the Laplace and osmotic pressures between the W 1 phase and W 2 phase is obtained. In addition, the W 1 /O/W 2 emulsion droplets are negatively charged, and the emulsion aggregation appears to be inhibited by electrostatic repulsion. More importantly, the Span 80 emulsifier at the O-W 2 interface seems to facilitate hydrogen bond formation between the hydrophilic moieties of Span 80 and the oxygen containing functional groups at the coal surface, which is verified by the results from molecular dynamics simulation (MDS). The hydrogen bond formation plays a leading role in the attachment and/or spreading of the emulsion droplets at the oxidized coal surface. The HIP W/O emulsion could be a potential economical and efficient replacement for the traditional oily collectors as a promising collector for the flotation of fine oxidized coal. [ABSTRACT FROM AUTHOR]

We describe a setup and procedures for contactless optical 3D-metrology of silicon micro-strip sensors. Space points are obtained by video microscopy and a high precision XY-table. The XY-dimensions are obtained from the movement of the table and pattern recognition, while the Z-dimension results from a Fast Fourier Transformation analyses of microscopic images taken at various distances of the optical system from the object under investigation, i.e. focus-based optical metrology. The setup is employed to measure the position of silicon sensors mounted onto a carbon fibre structure with a precision of a few microns. [ABSTRACT FROM AUTHOR]

Graphical abstract Highlights • Development of a magnetic trap to measure the torque exerted by braided DNA. • Detection of DNA torque from the dynamical response of superparamagnetic probes. • Description of DNA braiding dynamics using the model of a non-uniform oscillator. • DNA torque estimations from angular oscillations of superparamagnetic probes. Abstract We present a force spectroscopic method to quantify the torque exerted by braided DNA molecules. It involved a rotating magnetic field, with torsional stiffness comparable to the torque exerted by braided DNA molecules, to manipulate the macromolecules. During the initial stages of the braiding process, the probe rotated synchronously with the external field. However, when enough torsional free energy was accumulated in the molecules, the probe started oscillating. We exploited this oscillatory behavior for developing a procedure to estimate the torque exerted by the molecules. The procedure entailed modeling the system as a non-linear oscillator, which considered the dynamics of the angular orientation of the magnetic field vector with respect to the magnetic moment of the probe. [ABSTRACT FROM AUTHOR]

Graphical abstract Highlights • One Qing dynasty meticulous painting's pigments are non-destructively analyzed. • The painting was made no earlier than 1828 CE. • Brass powder has been used in Chinese painting in Qing dynasty. Abstract Three-dimensional video microscopy, Raman microscopy, and energy-dispersive X-ray fluorescence spectroscopy were applied on a Qing dynasty meticulous painting named 'Guan Gong Reads the Spring and Autumn Annals at Night'. All pigments were identified, including calcite, Paris green, hematite, gypsum, cinnabar, carbon black, quartz crystal, synthetic ultramarine blue and brass powder. Colors on the painting, in most parts, can be explained by a single pigment; while it is also true that some are caused by mixed pigments. The manufacture date of the painting should be no earlier than 1828. In addition, brass powder was used as pigment no later than the Qing dynasty. Finally, the combination of multiple non-destructive methods is an ideal for researching Chinese ancient paintings. [ABSTRACT FROM AUTHOR]

[Display omitted] • Classification, performance, function. • Characterisation techniques. • Future perspective. The significance of frothers in the context of mineral flotation is widely recognized, particularly in relation to their impact on bubble size and dispersion, coalescence inhibition, as well as the structure and stability of the froth. All these parameters hold substantial importance in determining the feasibility of the process including the overall recovery of valuable minerals and concentrate grade in flotation process. This review aims to offer a comprehensive overview of the importance of frothers in the flotation process focusing on their performance and function in the context of their chemical classification. The characterisation of flotation frothers is discussed, focusing on the most often used parameters including molecular weight, hydrophilic-lipophilic balance (HLB), dynamic frothability, and interfacial tension measurement. Additionally, the importance of advanced characterisation techniques such as single rising bubble experiments, high-speed video microscopy and infrared reflection absorption spectroscopy (IRRAS) is highlighted. Finally, the application of biodegradable and environmentally friendly biosurfactants such as rhamnolipids as "green frothers" in enhancing flotation rates is outlined. [ABSTRACT FROM AUTHOR]

The growing need for personalized, accurate, and non-invasive diagnostic technology has resulted in significant advancements, from pushing current mechanistic limitations to innovative modality developments across various disease-related biomarkers. However, there still lacks clinical solutions for analyzing multiple biomarkers simultaneously, limiting prognosis for patients suffering with complicated diseases or comorbidities. Here, we conceived, fabricated, and validated a multifrequency impedance cytometry apparatus with novel frequency-sensitive barcoded metal oxide Janus particles (MOJPs) as cell-receptor targeting agents. These microparticles are modulated by a metal oxide semi-coating which exhibit electrical property changes in a multifrequency electric field and are functionalized to target CD11b and CD66b membrane proteins on neutrophils. A multi-modal system utilizing supervised machine learning and simultaneous high-speed video microscopy classifies immune-specific surface receptors targeted by MOJPs as they form neutrophil-MOJP conjugates, based on multivariate multifrequency electrical recordings. High precision and sensitivity were determined based on the type of MOJPs conjugated with cells (>90% accuracy between neutrophil-MOJP conjugates versus cells alone). Remarkably, the design could differentiate the number of MOJPs conjugated per cell within the same MOJP class (>80% accuracy); which also improved comparing electrical responses across different MOJP types (>75% accuracy) as well. Such trends were consistent in individual blood samples and comparing consolidated data across multiple samples, demonstrating design robustness. The configuration may further expand to include more MOJP types targeting critical biomarker receptors in one sample and increase the modality's multiplexing potential. [ABSTRACT FROM AUTHOR]

Abstract Characterizing protein aggregates in the presence of silicone oil is a long standing challenge for the pharmaceutical industry. Silicone oil is often used as a lubricant in devices that deliver and store therapeutic protein products and has been linked to protein aggregation, which can compromise a drug's effectiveness or cause autoimmune responses in patients. Most traditional technologies cannot quantitatively distinguish protein aggregates and silicone oil in their native formulations for sizes less than 5 μm. We use holographic video microscopy to study protein aggregation to demonstrate its capability to quantitatively distinguish protein aggregates and silicone oil in the presence of varying amounts of the surfactants SDS and polysorbate 80 in the size range of 0.5-10 μm without the need for dilution or special sample preparation. We show that SDS denatures proteins and stabilizes silicone oil. We also show that polysorbate 80 may limit protein aggregate formation if it is added to an IgG solution before introducing silicone oil. [ABSTRACT FROM AUTHOR]

Video microscopy is one of the most versatile and successful measurement techniques in the spatio-temporal characterization of particle manipulations. The orientations of the imaging axes are always manually aligned correspondingly to the axes of the optical traps. We assume a residual misalignment angle as 5° exists between one imaging axis and the optical axis. Generous repeated position fluctuations resulted from Brownian motion are simulated, and the statistical results demonstrate dramatic cross-coupling effects occur in the situation of inconsistent trapping stiffness in orthogonal axes. The coupling effects may cause biased stiffness estimations and deteriorate the accuracy of parameter measurements related to position observations. For misalignment calibrations, multi-position observation methods using power imbalance and optical binding interactions are proposed with detailed discussions. The calibration can help to improve the accuracy of position related parameter measurements using suspended particles. [ABSTRACT FROM AUTHOR]

Most MUC5B mucin polymers in the upper airways of humans and pigs are produced by submucosal glands. MUC5B forms N-terminal covalent dimers that are further packed into larger assemblies because of low pH and high Ca2+ in the secretory granule of the mucin-producing cell. We purified the recombinant MUC5B N-terminal covalent dimer and used single- particle electron microscopy to study its structure under intracellular conditions. We found that, at intragranular pH, the dimeric MUC5B organized into head-to-head noncovalent tetramers where the von Willebrand D1-D2 domains hooked into each other. These N-terminal tetramers further formed long linear complexes from which, we suggest, the mucin domains and their C termini project radially outwards. Using conventional and video microscopy, we observed that, upon secretion into the submucosal gland ducts, a flow of bicarbonate- rich fluid passes the mucin-secreting cells. We suggest that this unfolds and pulls out the MUC5B assemblies into long linear threads. These further assemble into thicker mucin bundles in the glandular ducts before emerging at the gland duct opening. We conclude that the combination of intracellular packing of the MUC5B mucin and the submucosal gland morphology creates an efficient machine for producing linear mucin bundles. [ABSTRACT FROM AUTHOR]

Spermiogram tests are currently performed in two ways; computer assisted and visual assessment. Computerized techniques are costly and parameter dependent. Therefore, it is not preferred in many laboratories. Analysis based on visual assessment technique is subjective. In this study, we proposed a new computerized approach in which data acquisition is performed as in visual assessment, but analysis is done by computer. This approach provides more generalized results, requires less parameters due to the usage of standard counting chambers as in visual assessment technique, and cheaper than the other computerized techniques. Proposed approach includes two modules; video stabilization and motile sperm detection module. Stabilization is a requirement because it is impossible to fix proposed approach in ocular part completely. Otherwise, vibration affects the detection process of motile sperms. In this respect, feature-matching video stabilization idea is firstly utilized within the video-microscopy concept. Several feature extraction techniques were tested to sustain more stable videos in stabilization module. Motile sperm detection algorithm is then adapted to evaluate the efficiency of video stabilization and analyzing part. In case of vibrated frame sequence, the detection of total motile sperms concludes immediate peak values while it is around average values in stable frames. Samples were also evaluated by an expert with the visual assessment technique. Comparative tables and figures emphasize that the proposed approach can be employed in laboratories due to the high correlation with the visual assessment results. [ABSTRACT FROM AUTHOR]

Primary ciliary dyskinesia (PCD) is a rare genetic disorder causing a defective ciliary structure, which predominantly leads to an impaired mucociliary clearance and associated airway disease. As there is currently no single diagnostic gold standard test, PCD is diagnosed by a combination of several methods comprising genetic testing and the examination of the ciliary structure and function. Among the approved diagnostic methods, only high-speed video microscopy (HSVM) allows to directly observe the ciliary motion and therefore, to directly assess ciliary function. In the present work, we present our recently developed freely available open-source software – termed "Cilialyzer", which has been specifically designed to support and facilitate the analysis of the mucociliary activity in respiratory epithelial cells captured by high-speed video microscopy. In its current state, the Cilialyzer software enables clinical PCD analysts to load, preprocess and replay recorded image sequences as well as videos with a feature-rich replaying module facilitating the commonly performed qualitative visual assessment of ciliary function (including the assessment of the ciliary beat pattern). The image processing methods made accessible through an intuitive user interface allow clinical specialists to comfortably compute the ciliary beating frequency (CBF), the activity map and the "frequency correlation length" – an observable getting newly introduced. Furthermore, the Cilialyzer contains a simple-to-use particle tracking interface to determine the mucociliary transport speed. Cilialyzer is fully written in the Python programming language and freely available under the terms of the MIT license. The proper functioning of the computational analysis methods constituting the Cilialyzer software is demonstrated by using simulated and representative sample data from clinical practice. Additionally, the software was used to analyze high-speed videos showing samples obtained from healthy controls and genetically confirmed PCD cases (DNAI1 and DNAH11 mutations) to show its clinical applicability. Cilialyzer serves as a useful clinical tool for PCD analysts and provides new quantitative information awaiting to be clinically evaluated using cohorts of PCD. As Cilialyzer is freely available under the terms of a permissive open-source license, it serves as a ground frame for further development of computational methods aiming at the quantification and automation of the analysis of mucociliary activity captured by HSVM. • The Cilialyzer is an open-source software serving as a useful clinical tool. • It facilitates and improves the clinical examination of ciliary function in PCD suspects. • Comfortable computation of quantitative information – such as the CBF and MCC speed. • Presents a first big step towards a quantitative HSVM analysis of ciliary function. [ABSTRACT FROM AUTHOR]

The accurate analysis of microscopy images representing various materials obtained in scanning probe microscopy, scanning tunneling microscopy, and transmission electron microscopy, is in general time consuming as it requires the inspection of multiple data bases for the correct interpretation of the observed crystal structures. This task is especially demanding in microscopy video analysis involving a vast amount of image data. The recent development of deep learning (DL) algorithms has paved the way for cutting-edge microscopy studies in materials science, often outperforming conventional image analysis methods. This paper reviews the state-of-the-art in DL-based synthetic data generation, materials structure identification, three-dimensional structural reconstruction, and physical properties evaluation for different types of microscopy images. First, the fundamental concepts of DL relevant to materials science applications are reviewed. Subsequently, the combined experimental measurements and numerical simulations for preparing dedicated microscopy image for DL analysis are discussed. Then, the review concentrates on the core topic of the paper, that is the critical assessment of DL advances in materials' structural and physical properties evaluation. We believe that the future development and deployment of DL for practical microscopy data analysis will rely on the progress and improvement of advanced algorithms and innovative methods for training data generation. [ABSTRACT FROM AUTHOR]

Oil–water dispersed flows produced at valves or restrictions are very common in industry. For example, crude oil desalting processes normally use dispersion valves to mix the dilution water. In this case, the knowledge of dispersed droplet sizes is crucial for the design and optimum control of the process. However, little work has been devoted to characterize and model droplet sizes produced in industrial-type valves. The present work studies water droplet sizes produced by the passage of oil–water flow through a globe valve mounted in a large scale flow loop of 0.1 m internal diameter. Experiments were performed under different pressure drops across the globe valve, and different oil and water flow rates. Produced water droplet sizes were measured in situ downstream from the globe valve location using particle video microscopy. Droplet sizes were compared with theoretical scales for turbulent and viscous break-up. The effect of the volume fraction of dispersed phase on droplet size was also examined. In addition, droplet size distributions were analyzed and fitted using known statistical functions. [ABSTRACT FROM AUTHOR]

We present an improved version of the magnetic tweezers/viscous drag (MTVD) hybrid assay that allows dynamic torque measurements during the spontaneous relaxation of DNA braids. The method relies on the ability of pairs of DNA molecules to rotate a microscopic probe attached to their upper ends using the elastic energy stored when braided. Digital tracking of the rotation of the dumbbell against the viscous drag allowed estimation of the torque exerted by the molecules. The unwinding dynamics of DNA braids against different amounts of viscous drag was characterized employing probes of two different sizes. We identified and characterized different perturbations that may affect the unbraiding dynamics. Estimates of the torques associated to the process of DNA unbraiding are reported. [ABSTRACT FROM AUTHOR]

• In situ video microscopy allows studying of microneedle growth in a miniature cell. • Different longitudinal and lateral growth mechanisms can occur simultaneously. • Microneedles have a carcass/filler structure; carcass consists of core and branches. • Needle surface structure is either highly regular or irregular; tips are also varied. Composite germanium/germanium oxide needles with a carcass/filler structure, thicknesses of 0.25–10 µm and lengths up to 450 µm were grown by chemical transport involving gaseous GeO. The synthesis was carried out at temperatures of 897–1100 K in a newly developed miniature cell. Through a top window one can observe the growth process in situ via video microscopy. The cell can be evacuated and sealed, enclosing all products generated in reactions, but can easily be opened afterwards for an ex situ analysis of the needles. The in situ observations show that the needles grow with longitudinal velocities of up to 30 µm/s, depending on temperature and needle thickness, and with lateral velocities of up to 0.11 µm/s. The surface structure and the cross-sections of the needles were studied ex situ by SEM as well as EDX spectroscopy combined with element mapping. The surface structure of needles exhibits highly regular arrays of germanium/germanium oxide microunits but also shows irregularities and defects. The study of the longitudinal section and the lamella showed that this regularity is remarkable also inside the needle all way down to the core. Furthermore, according to the SAED and Raman spectroscopy, the needle carcass (core with branches) is mainly composed of a cubic Ge and an amorphous filling mass of Ge and O, which presumably corresponds to GeO 2. [ABSTRACT FROM AUTHOR]

Despite its importance, little attention has been paid to the effect of freezing on starch gelatinization during subsequent heating. In an effort to address this problem, the effects of freezing and water accessibility in starch gelatinization during heating were analyzed in situ using hot-stage polarized-light video-microscopy. Native starch granules immersed in water, carrageenan gel or sucrose solution were heated at 15 °C/min or frozen for 48 h and heated at 15 °C/min. Energy changes were measured using differential scanning calorimetry. Surface morphology was examined using scanning electron microscopy and atomic force microscopy. The results showed that freezing delayed the gelatinization degree and that the addition of carrageenan delayed the gelatinization process. That process was further delayed when sucrose was added. The enthalpy of gelatinization decreased accordingly (ΔHstarch -water-sucrose > ΔHstarch-water-carrageenan > ΔHstarch -Water)- The temperature range over which the degree of gelatinization differed between frozen and unfrozen samples was diminished when accessibility to water was reduced. [ABSTRACT FROM AUTHOR]

A rapidly increasing number of papers describing novel iPSC models for cardiac diseases are being published. To be able to understand the disease mechanisms in more detail, we should also take the full advantage of the various methods for analyzing these cell models. The traditionally and commonly used electrophysiological analysis methods have been recently accompanied by novel approaches for analyzing the mechanical beatingbehavior of the cardiomyocytes. In this review, we provide first a concise overview on the methodology for cardiomyocyte functional analysis and then concentrate on the video microscopy, which provides a promise for a new faster yet reliable method for cardiomyocyte functional analysis. We also show how analysis conditions may affect the results. Development of the methodology not only serves the basic research on the disease models, but could also provide the much needed efficient early phase screening method for cardiac safety toxicology. This article is part of a Special Issue entitled: Cardiomyocyte Biology: Integration of Developmental and Environmental Cues in the Heart edited by Marcus Schaub and Hughes Abriel. [ABSTRACT FROM AUTHOR]

• Millimetres long bundles of hydrate fibres grow from surface macropores of a porous substrate. • Surface macropores confirmed by confocal reflectance and scanning electron microscopies. • X-ray radiography shows fluid exchange in the peripheral layers of the substrate. • X-ray computed microtomography reveals a belt of organised peripheral porosity and connection to surface macropores. • These features explain the unprecedented observation of comb or whale-bone like bundles of hydrate fibres. Venet et al., reported unprecedented, bundled growth of ≈ 1-10 mm long, hollow gas hydrate fibres, diameter ≈ 1-10 μ m, on an activated carbon close to a guest-water interface, circumventing the hydrate growth bottleneck. Here, in situ video microscopy traces growth (2-3 μ m s - 1 at 0.5 ° C) to surface macropores of similar diameter, further exhibited by confocal and scanning electron microscopies. X-ray radiography shows fluid exchange to a depth of ≈ 50-100 μ m in a total porous fraction 40 % (gravimetric porometry). Computed X-ray microtomography reveals a belt of organised peripheral porosity of aligned ≈ 5 μ m pores. These structures explain the origin of the comb or whalebone-like appearance of the hydrate. Whereas hydrate promotion by porous media is usually assumed to proceed inside the materials, like tiny hydrate storage warehouses, we find that the substrate behaves more like a 'lean' or 'just in time' production line, with hydrate extruded as fast as it is produced near the surface. [ABSTRACT FROM AUTHOR]

Aside from nerve agents, various highly toxic pesticides belong to the group of organophosphorus (OP) compounds, thereby causing a large number of intoxications every year. Unfortunately, there are still shortcomings in the current treatment for OP poisoning and research on novel therapeutic options is restricted in several aspects. In this study we investigated the suitability of organotypic cocultures for pharmacological in vitro studies involving OP compounds. These slice cultures are derived from murine spinal cord and muscle tissue forming functional neuromuscular synapses, which trigger spontaneous contractions of muscle fibers. Using video microscopy to quantify muscle activity, we assessed the viability of cocultures after exposure to soman and VX, and the associated loss and recovery of neuromuscular function. Antidotal treatment was not provided. The application of nerve agents led to an almost complete loss of muscle activity. However, cell cultures regained equivalent muscular function to the control situation three and seven days after intoxication. In summary, the tested in vitro system could be a promising tool for the investigation of long term effects and therapeutic options for OP poisoning. [ABSTRACT FROM AUTHOR]

Juglone 1 an plumbagin 2 are plant secondary metabolites nowadays well known for their anticancer properties. In this study we synthesized analogues of 1 and 2 deriving from the functionalization of the OH group in position 5 with different side chains in form of esters and ethers. Therefore the growth inhibitory activities of these adducts were evaluated in vitro on six cancer cell lines using the MTT colorimetric assays along with the two natural parent compounds. The data revealed that these latter displayed the strongest growth inhibitory activities in vitro. Quantitative videomicroscopy analyses were then carried out on human U373 glioblastoma cells, which are characterized by various level of resistance to pro-apoptotic stimuli. We compared the naturally occurring reference compounds 1 and 2 with the derivatives exerting the best activities in terms of IC 50 growth inhibitory values. These analyses showed that both juglone and plumbagin had a cytostatic effect on U373 cells and were able to overcome the intrinsic resistance of U373 cancer cells to pro-apoptotic stimuli. [ABSTRACT FROM AUTHOR]

The purpose of this study was to determine if the cardiovascular response to hypoxia was altered by the presence of bisphenol A (BPA) in Danio rerio embryos. It was expected that BPA exposure would affect cardiovascular parameters during hypoxia more than normoxia due to an interaction between BPA and the hypoxia-inducible factor (HIF-1α) pathway. We demonstrate that BPA exposure has a minimal effect during normoxia but can severely affect the cardiovascular system during a hypoxic event. Cardiovascular response was measured in vivo using video microscopy and digital motion analysis. RBC density increased 35% in hypoxia alone but decreased 48% with addition of 0.25 mg/L BPA. Tissue vascularization (% coverage) was unaffected by hypoxia alone but decreased 37% with addition of 0.25 mg/L BPA. The diameter and RBC velocity of arteries were more sensitive than veins to BPA exposure during both normoxia and hypoxia. Arterial RBC velocity decreased 42% during normoxia and 52% during hypoxia with 1 mg/L BPA. This decrease in velocity may in part be due to the 86% decrease in heart rate (ƒ H ) observed during co-exposure to hypoxia and 5 mg/L BPA. While stroke volume (S V ) was unaffected by treatment, cardiac output (Q) decreased by 69% with co-exposure. ƒ H and Q were not affected by BPA exposure during normoxia. Development ultimately slowed by 146% and mortality rates were 95% during hypoxia when exposed to 5 mg/L BPA. Our results show for the first time that BPA exposure alters the cardiovascular system during hypoxia more so than normoxia. [ABSTRACT FROM AUTHOR]

The single droplet freezing apparatus described by Pore et al. (2009), which allows crystallisation to be monitored in situ by X-ray diffraction, was modified to allow rapid switching of coolant gas and monitoring by video microscopy. The apparatus was used to study drops of cocoa butter undergoing simulated spray freezing at high cooling rates, e.g . 130 K/min. The transformation of an Ivory Coast cocoa butter to the Form V polymorph was significantly faster in drops (∼40 h) than in static bulk samples (10 days) crystallised under isothermal conditions. Phase transformation was observed from Forms I/II → III → IV → melt → V, with Form V crystallising directly from the melt at 28.6 °C. Numerical simulations of the temperature evolution within the droplet established that the drops are not isothermal, explaining why nucleation was initially observed in the lower (upstream) part of the droplet. [ABSTRACT FROM AUTHOR]

In the present study, for the first time in fish spermatozoa, we describe the precise chronology of motility initiation of sterlet (sturgeon) sperm from completely immotile flagella to regular full wave propagation. The successive activation steps were investigated by high-speed video microscopy, using specific experimental situation, where sperm motility initiation was delayed in time up to several seconds (10 ± 2.68 seconds). Starting from fully immotile, the flagellum shows some trembling for a brief period, soon followed by appearance of the first real bend (so-called “principal bend”) with a large wave amplitude 4.28 ± 0.65 μm, then by the “reverse bend,” the latter presenting a lower (P < 0.05) wave amplitude (1.14 ± 0.32 μm). This couple of first bends formed at the basal region begins to propagate toward the flagellar tip but gradually fades when reaching the midflagellum, wherein consequently the sperm cell remains nonprogressive. This behavior repeats several times until a stage where the amplitude of the reverse bend gradually reaches a value similar that of the principal bend: The larger amplitude of this couple of bends finally leads to sustain a real “takeoff” of the sperm cell characterized by a full flagellar wave propagation generating an active forward displacement similar to that occurring during regular steady state motility (several seconds after activation). Starting from the earliest stages of motility initiation, the wave propagation along the flagellum and formation of new waves proceeded in a helical manner leading to a 3-dimensional rotation of the whole spermatozoon. Eventually, we estimated that the time period needed from the activation signal (contact with fresh water) to full wave propagation ranges from 0.4 to 1.2 seconds. [ABSTRACT FROM AUTHOR]

Effects of applied potential on stable pitting corrosion of 304 stainless steel were studied by potentiostatic polarization and three-dimensional video microscope. The results showed that pit mouth morphologies and three-dimensional structures were dependent on potential. Higher potentials resulted in more open pits. Pit volume, depth, and width increased with potential. The ratio of pit depth to width continuously changed with time and potential. The pitting current density of a single pit increased with potential, indicating that an ohmic potential drop presented between the pit internal surface and the bulk solution, but stable propagation was still controlled by corrosion products diffusion. [ABSTRACT FROM AUTHOR]

Excess sodium as salt (NaCl) added to foods may be harmful to humans. Most salt intake comes from processed foods, therefore salt reduction strategies should be developed by the food industry. One strategy seeks to optimize the delivery of salt ions inside the mouth. To accomplish this purpose, the dissolution characteristics of salt crystals in saliva must be understood. However, relatively scarce information is available on this topic. This study compares the dissolution of five commercial salts in deionized water and two formulations of artificial saliva, at four temperatures and in the absence of agitation. Salt dissolution was quantified in vitro by analysis of video-microscopy images taken at different times. Higher dissolution rates of salt crystals were found in water than in artificial saliva, and at higher temperatures. Video-microscopy was instrumental to reveal that some crystals were fragmented during dissolution while others remained as a unit of decreasing size until complete solubilization. Increased surface area after fragmentation led to pyramid-shape crystals having the highest dissolution rates. Hence, significant changes can be achieved in the dissolution of salt depending on the crystal structure and its dissolution pattern. [ABSTRACT FROM AUTHOR]

Label free time-lapse video microscopy based monitoring of time evolving cell population morphology has potential to offer a simple and cost effective method for identity control of cell lines. Such morphology monitoring also has potential to offer discovery of chemically induced differential changes between pairs of cell lines of interest, for example where one in a pair of cell lines is normal/sensitive and the other malignant/resistant. A new simple algorithm, pixel histogram hierarchy comparison (PHHC), for comparison of time evolving morphologies (TEM) in phase contrast time-lapse microscopy movies was applied to a set of 10 different cell lines and three different iso-genic colon cancer cell line pairs, each pair being genetically identical except for a single mutation. PHHC quantifies differences in morphology by comparing pixel histogram intensities at six different resolutions. Unsupervised clustering and machine learning based classification methods were found to accurately identify cell lines, including their respective iso-genic variants, through time-evolving morphology. Using this experimental setting, drugs with differential activity in iso-genic cell line pairs were likewise identified. Thus, this is a cost effective and expedient alternative to conventional molecular profiling techniques and might be useful as part of the quality control in research incorporating cell line models, e.g. in any cell/tumor biology or toxicology project involving drug/agent differential activity in pairs of cell line models. [ABSTRACT FROM AUTHOR]

Abstract: Flow quantification with high-frequency (>20 MHz) power Doppler ultrasound can be performed objectively using the wall-filter selection curve (WFSC) method to select the cutoff velocity that yields a best-estimate color pixel density (CPD). An in vivo video microscopy system (IVVM) is combined with high-frequency power Doppler ultrasound to provide a method for validation of CPD measurements based on WFSCs in mouse testicular vessels. The ultrasound and IVVM systems are instrumented so that the mouse remains on the same imaging platform when switching between the two modalities. In vivo video microscopy provides gold-standard measurements of vascular diameter to validate power Doppler CPD estimates. Measurements in four image planes from three mice exhibit wide variation in the optimal cutoff velocity and indicate that a predetermined cutoff velocity setting can introduce significant errors in studies intended to quantify vascularity. Consistent with previously published flow-phantom data, in vivo WFSCs exhibited three characteristic regions and detectable plateaus. Selection of a cutoff velocity at the right end of the plateau yielded a CPD close to the gold-standard vascular volume fraction estimated using IVVM. An investigator can implement the WFSC method to help adapt cutoff velocity to current blood flow conditions and thereby improve the accuracy of power Doppler for quantitative microvascular imaging. [Copyright &y& Elsevier]

Abstract: This study applies 2-D speckle tracking using B-scan ultrasound imaging to estimate the instantaneous and total displacement of the middle flexor digitorum superficialis (FDS) tendon proximal to the wrist. This is achieved by performing the study with human patients, during regular carpal tunnel surgeries. B-Scan images were collected with a 12-MHz transducer placed proximal to the wrist, while a video microscope simultaneously imaged the exposed flexor tendons in the palm as a reference for validation. The accuracy of the proposed speckle-based tracking method is compared using log-compressed Rayleigh (Fisher-Tippet)-, Gaussian (sum of squared differences)- and Laplacian (sum of absolute differences)-based statistics as similarity measures. Overall, tracking was successful and the Rayleigh technique performed better than the Laplacian or Gaussian technique. One goal of this research was to non-invasively monitor FDS tendon displacement in the wrist for the purposes of controlling a prosthetic device. An additional goal was to obtain pre- and post-operative clinical information. [Copyright &y& Elsevier]

Abstract: Background: In recent years, a number of tissue factor pathway inhibitor (TFPI) antagonists have been developed to serve as bypassing agents to improve hemostasis in hemophilia A. Since TFPI antagonists and FVIII concentrates are procoagulants, their combined effect on spatial clot formation could be potentially pro-thrombotic. Objective: To investigate the cooperative effect of TFPI inhibition and supplementation of FVIII in hemophilia A in a spatial, reaction-diffusion experiment in vitro. Methods: Plasma was collected at different time points from hemophilia A patients undergoing prophylaxis and was supplemented in vitro with TFPI inhibitor BAX499 (formerly ARC19499) at concentrations from 0 up to 600nM. Clotting propagation in recalcified plasma activated by a surface with immobilized tissue factor (TF) was monitored by videomicroscopy. Results: Increasing concentration of BAX499 improved coagulation for all hemophilia A plasma samples activated with TF at 1.6pmole/m2 by shortening lag time and increasing initial clot growth velocity and clot size. In contrast, plasma concentration of FVIII had little effect on lag time, but increased spatial clot growth velocity. There was a decrease in the BAX499 efficiency as FVIII concentration increased (lag time shortened by 50% if FVIII:C<5%, but the effect was only 25% if FVIII:C>30%). Conclusions: The results indicate that BAX499 has an effect on clotting in hemophilia A plasma at low FVIII concentrations, however has little effect at high FVIII concentrations. [Copyright &y& Elsevier]

Automated visual tracking of cells from video microscopy has many important biomedical applications. In this paper, we track human monocyte cells in a fluorescent microscopic video using matching and linking of bipartite graphs. Tracking of cells over a pair of frames is modeled as a maximum cardinality minimum weight matching problem for a bipartite graph with a novel cost function. The tracking results are further refined using a rank-based filtering mechanism. Linking of cell trajectories over different frames are achieved through composition of bipartite matches. The proposed solution does not require any explicit motion model, is highly scalable, and, can effectively handle the entry and exit of cells. Our tracking accuracy of (97.97±0.94)% is superior than several existing methods [(95.66±2.39)% [11], (94.42±2.08)% [20], (81.22±5.75)% [13], (78.31±4.70)% [14]] and is highly comparable (98.20±1.22)% to a recently published algorithm [26]. [ABSTRACT FROM AUTHOR]

Abstract: Particle tracking microrheology, an emerging experimental technique, which utilizes the Brownian motion of embedded particles to probe local dynamics of soft materials, is presented. Particle tracking microrheology is a powerful technique that enables the measurement of viscoelastic responses in small sample volumes, which are inaccessible to macrorheology and to spatially map structural heterogeneities at a microlevel. Therefore, particle tracking microrheology has considerable potential in food emulsions and gels, since these systems are commonly inhomogeneous. Recent advances and achievements are discussed, including the basic principles, operating regimes and limitations of the technique. The application of the technique in the field of food gels and emulsions to study the evolving dynamics of inhomogeneous at microscale length systems and during sol–gel transition is highlighted. [Copyright &y& Elsevier]