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Urban growth is typically considered a process of expansion. As population grows and transport costs decrease urban density gradients are expected to gradually flatten. This is a basic feature of cities, explained by urban economic models and empirically supported by a plethora of studies about urban density development from all over the world. However, additional forces, such as changes in demographic composition and locational preferences of the urban population acting at local levels, may counteract the flattening tendency of urban gradients. In this paper, we suggest a methodology to test the impact of local density changes on urban gradients, looking at spatiotemporal developments in terms of housing and population. Using highly detailed data on individual housing units and inhabitants in major Dutch cities, we first assess and compare urban density gradients during the period 2000–2017. In all the analysed Dutch cities, both dwelling and population density gradients are becoming steeper over time, contradicting standard predictions from urban economic literature and empirical reports worldwide. The observed trend of steepening urban gradients is partly explained by the presence of historical monuments and urban amenities. 城市发展通常被认为是一个扩张的过程。随着人口增长和运输成本降低，城市密度梯度 预计将逐渐扁平。这是城市的一个基本特征，由城市经济模型解释，并由世界各地关于 城市密度发展的大量研宄实证支持。然而，额外的力量，如在地方一级行动的城市人口 的人口构成和位置偏好的变化，可能抵消城市梯度的扁平趋势。在这篇文章中，我们提 出了一种方法来测试局部密度变化对城市梯度的影响，即从住房和人口的角度来看时空 发展。我们首先使用荷兰主要城市的个人住房单元和居民的非常详细的数据来评估和比 较 2000_2017年期间的城市密度梯度。在所有被分析的荷兰城市中，随着时间的推移，居住和人 口密度梯度都变得越来越陡，这与世界范围内城市经济文献和经验报告的标准预测相矛 盾。观察到的城市梯度变陡的趋势部分是由历史遗迹和城市便利设施的存在来解释的。

Virus purification by cesium chloride (CsCl) density gradient, which generally requires an expensive ultracentrifuge, is an essential technique in virology. Here, we optimized virus purification by CsCl density gradient using general centrifugation (40,000 × g , 2 h, 4 °C), which showed almost the same purification ability as conventional CsCl density gradient ultracentrifugation (100,000 × g , 1 h, 4 °C) using phages S13′ and φEF24C. Moreover, adenovirus strain JM1/1 was also successfully purified by this method. We suggest that general centrifugation can become a less costly alternative to ultracentrifugation for virus purification by CsCl densiy gradient and will thus encourage research in virology.

Lipoprotein lipase (LPL), the enzyme that hydrolyzes triglycerides in plasma lipoproteins, is assumed to be active only as a homodimer. In support of this idea, several groups have reported that the size of LPL, as measured by density gradient ultracentrifugation, is ∼110 kDa, twice the size of LPL monomers (∼55 kDa). Of note, however, in those studies the LPL had been incubated with heparin, a polyanionic substance that binds and stabilizes LPL. Here we revisited the assumption that LPL is active only as a homodimer. When freshly secreted human LPL (or purified preparations of LPL) was subjected to density gradient ultracentrifugation (in the absence of heparin), LPL mass and activity peaks exhibited the size expected of monomers (near the 66-kDa albumin standard). GPIHBP1-bound LPL also exhibited the size expected for a monomer. In the presence of heparin, LPL size increased, overlapping with a 97.2-kDa standard. We also used density gradient ultracentrifugation to characterize the LPL within the high-salt and low-salt peaks from a heparin-Sepharose column. The catalytically active LPL within the high-salt peak exhibited the size of monomers, whereas most of the inactive LPL in the low-salt peak was at the bottom of the tube (in aggregates). Consistent with those findings, the LPL in the low-salt peak, but not that in the high-salt peak, was easily detectable with single mAb sandwich ELISAs, in which LPL is captured and detected with the same antibody. We conclude that catalytically active LPL can exist in a monomeric state.

Energetic electron precipitation from the equatorial magnetosphere into the atmosphere plays an important role in magnetosphere‐ionosphere coupling: precipitating electrons alter ionospheric properties, whereas ionospheric outflows modify equatorial plasma conditions affecting electromagnetic wave generation and energetic electron scattering. However, ionospheric measurements cannot be directly related to wave and energetic electron properties measured by high‐altitude, near‐equatorial spacecraft, due to large mapping uncertainties. We aim to resolve this by projecting low‐altitude measurements of energetic electron precipitation by ELFIN CubeSats onto total electron content (TEC) maps serving as a proxy for ionospheric density structures. We examine three types of precipitation on the nightside: precipitation of <200 keV electrons in the plasma sheet, bursty precipitation of <500 keV electrons by whistler‐mode waves, and relativistic (>500 keV) electron precipitation by EMIC waves. All three types of precipitation show distinct features in TEC horizontal gradients, and we discuss possible implications of these features. Plain Language Summary Bursty precipitation of energetic electrons, via pitch‐angle scattering by whistler‐mode waves from the magnetosphere to the ionosphere, is an important factor in the global magnetosphere‐ionosphere coupling. It induces local modifications of ionospheric density and chemical composition. A recurrent problem in the investigation of this process is the presence of large uncertainties in the field‐line mapping between ionospheric density structures and high altitude satellites measuring electron fluxes in the magnetosphere. In the present study, such uncertainties are significantly reduced by making use of precipitating electron fluxes recorded by ELFIN CubeSats at low altitudes (450 km) just above the ionosphere and comparing them with maps of the corresponding ionospheric density structures. We identify three different types of electron precipitation on the nightside, corresponding to low, moderate, and high energy precipitating electrons. We show that each type of the precipitation is characterized by particular plasma density gradients in the ionosphere, suggesting a key role of wave ducting by plasma density gradients in fostering the precipitation of 300–500 keV electrons by whistler‐mode waves, and the potential importance of midnight plasma injections in generating EMIC waves that can further precipitate 0.5–2 MeV electrons far away from the plasmasphere. Key Points Total electron content (TEC) maps are examined during energetic electron precipitation at the conjugate low altitude Night‐side whistler‐mode wave driven precipitation is often associated with local horizontal TEC gradients Night‐side EMIC wave driven precipitation is often poleward of the TEC minima associated with the plasmapause projection

Studying neutrophils is challenging due to their limited lifespan, inability to proliferate, and resistance to genetic manipulation. Neutrophils can sense various cues, making them susceptible to activation by blood collection techniques, storage conditions, RBC lysis, and the isolation procedure itself. Here we assessed the impact of the five most used methods for neutrophil isolation on neutrophil yield, purity, activation status and responsiveness. We monitored surface markers, reactive oxygen species production, and DNA release as a surrogate for neutrophil extracellular trap (NET) formation. Our results show that neutrophils isolated by negative immunomagnetic selection and density gradient methods, without RBC lysis, resembled untouched neutrophils in whole blood. They were also less activated and more responsive to milder stimuli in functional assays compared to neutrophils obtained using density gradients requiring RBC lysis. Our study highlights the importance of selecting the appropriate method for studying neutrophils, and underscores the need for standardizing isolation protocols to facilitate neutrophil subset characterization and inter-study comparisons.

As a new functional graded lattice structure construction strategy, the relative density gradient strategy has a promising future due to its ease of realization in various lattice structures. This paper proposes a BCC lattice structure combining two different lattice single cells. Based on this, the single cells of different structures are assigned different relative density gradients, resulting in 18 combined gradient lattice structures. Based on proving the experimental feasibility of numerical simulation, the mechanical properties and energy absorption characteristics of the combined gradient lattice structure are investigated by numerical simulation. When applied to composite lattice structures, the proposed wave-like gradient design significantly improves mechanical properties. Among the various gradient strategies examined, several have achieved mechanical performance close to that of uniform lattice structures. To some extent, this approach mitigates the common drawback of gradient lattice structures—where the relative density of the weakest layer is consistently lower than the interlayer relative density of uniform lattice structures—resulting in varying degrees of mechanical performance degradation compared to their uniform counterparts. The proposed linearly enhanced gradient strategy (Strategy-LE) possesses higher SEA and CLE values when the lattice structure is subjected to compressive loading, with an improvement of 6.36% in SEA and 61.6% in CLE over the uniform structure. Through the relative density gradient design, the adaptability of the BCC lattice structure in actual complex application scenarios is greatly enhanced, and the energy-absorbing properties of the lattice structure are greatly improved.

Ultracentrifugation (UC) is recognized as a robust approach for the isolation of extracellular vesicles (EVs). However, recent studies have highlighted limitations of UC including low recovery efficiencies and aggregation of EVs that could impact downstream functional analyses. We tested the benefit of using a liquid cushion of iodixanol during UC to address such shortcomings. In this study, we compared the yield and purity of EVs isolated from J774A.1 macrophage conditioned media by conventional UC and cushioned-UC (C-UC). We extended our study to include two other common EV isolation approaches: ultrafiltration (UF) and polyethylene glycol (PEG) sedimentation. After concentrating EVs using these four methods, the concentrates underwent further purification by using OptiPrep density gradient ultracentrifugation (DGUC). Our data show that C-DGUC provides a two-fold improvement in EV recovery over conventional UC-DGUC. We also found that UF-DGUC retained ten-fold more protein while PEG-DGUC achieved similar performance in nanoparticle and protein recovery compared to C-DGUC. Regarding purity as assessed by nanoparticle to protein ratio, our data show that EVs isolated by UC-DGUC achieved the highest purity while C-DGUC and PEG-DGUC led to similarly pure preparations. Collectively, we demonstrate that the use of a high-density iodixanol cushion during the initial concentration step improves the yield of EVs derived from cell culture media compared to conventional UC. This enhanced yield without substantial retention of protein contaminants and without exposure to forces causing aggregation offers new opportunities for the isolation of EVs that can subsequently be used for functional studies.

Viral vector-based gene therapies and vaccines require accurate characterization of capsid species. The current gold standard for assessing capsid loading of adeno-associated virus (AAV) is sedimentation velocity analytical ultracentrifugation (SV-AUC). However, routine SV-AUC analysis is often size-limited, especially without the use of advanced techniques (e.g., gravitational-sweep) or when acquiring the multiwavelength data needed for assessing the loading fraction of viral vectors, and requires analysis by specialized software packages. Density gradient equilibrium AUC (DGE-AUC) is a highly simplified analytical method that provides high-resolution separation of biologics of different densities (e.g., empty and full viral capsids). The analysis required is significantly simpler than SV-AUC, and larger viral particles such as adenovirus (AdV) are amenable to characterization by DGE-AUC using cesium chloride gradients. This method provides high-resolution data with significantly less sample (estimated 56-fold improvement in sensitivity compared to SV-AUC). Multiwavelength analysis can also be used without compromising data quality. Finally, DGE-AUC is serotype-agnostic and amenable to intuitive interpretation and analysis (not requiring specialized AUC software). Here, we present suggestions for optimizing DGE-AUC methods and demonstrate a high-throughput AdV packaging analysis with the AUC, running as many as 21 samples in 80 min.

In the present study, a series of lattice structures with Gyroid minimal surfaces were meticulously designed to incorporate linear density gradients and two distinct trigonometric function-based density gradients. These advanced architectures were subsequently compared and contrasted with a uniform lattice sandwich structure. The mechanical behavior and energy absorption characteristics of the four lattice sandwich structures were rigorously investigated through a combination of experimental testing and finite element analysis (FEA). The results of this comprehensive analysis revealed that during compression, all four gradient lattice structures exhibited varying degrees of shear slip, which manifested as discernible discrepancies in their respective stress–strain curves. Relative to the uniform lattice structure, the linear gradient lattice sandwich structure exhibited an enhancement in elastic modulus by 1.69%, while the square sine function gradient lattice sandwich structure showed a significant increase of 14.45% in elastic modulus. Conversely, the square cosine function gradient lattice sandwich structure experienced a reduction in elastic modulus by 9.61%. Employing either a linear gradient or a square sine function density gradient design was found to augment the load-bearing capacity of the uniform lattice structure. Notably, when the strain in the uniform structure reached densification strain, it absorbed energy exceeding 5.842 MJ/m 3 , indicating superior energy absorption capabilities among the four structures examined, thus rendering it particularly suitable for applications where high energy absorption is imperative. Furthermore, finite element simulations were conducted to validate the experimental findings, and the simulation results demonstrated a high degree of correlation with the experimental data, with discrepancies less than 6%, thereby confirming the reliability of the FEA model in predicting the performance of these intricate lattice structures.

Bone marrow mononuclear cells (BMNCs) are widely used in regenerative medicine, but recent data suggests that the isolation of BMNCs by commonly used Ficoll-Paque density gradient centrifugation (DGC) causes significant cell loss and influences graft function. The objective of this study was to determine in an animal study whether and how Ficoll-Paque DGC affects the yield and composition of BMNCs compared to alternative isolation methods such as adjusted Percoll DGC or immunomagnetic separation of polymorphonuclear cells (PMNs). Each isolation procedure was confounded by a significant loss of BMNCs that was maximal after Ficoll-Paque DGC, moderate after adjusted Percoll DGC and least after immunomagnetic PMN depletion (25.6±5.8%, 51.5±2.3 and 72.3±6.7% recovery of total BMNCs in lysed bone marrow). Interestingly, proportions of BMNC subpopulations resembled those of lysed bone marrow indicating symmetric BMNC loss independent from the isolation protocol. Hematopoietic stem cell (HSC) content, determined by colony-forming units for granulocytes-macrophages (CFU-GM), was significantly reduced after Ficoll-Paque DGC compared to Percoll DGC and immunomagnetic PMN depletion. Finally, in a proof-of-concept study, we successfully applied the protocol for BMNC isolation by immunodepletion to fresh human bone marrow aspirates. Our findings indicate that the common method to isolate BMNCs in both preclinical and clinical research can be considerably improved by replacing Ficoll-Paque DGC with adapted Percoll DGC, or particularly by immunodepletion of PMNs.