Spontaneous Particle Ordering, Sorting, and Assembly on Soap Films.

Soap bubbles exhibit abundant fascinating phenomena throughout the entire life of evolution with different fundamental physics governing them. Nevertheless, the complicated dynamics of small objects in soap films are still unrevealed. Here, we report the first observation of spontaneous particle ordering in a complicated galaxy of soap films without any external energy. The balance of interfacial tension at two liquid-gas interfaces is theoretically predicted to govern belted wetted particles (BWPs) traveling along a specified path spontaneously. Such spontaneous particle path-finding is found to depend on the particle size and hydrophilic properties. Spontaneous particle sorting is directly realized via these discrete and distinctive paths for different particles. The deformation of the soap membrane facilitates 1D/2D particle organization along the path. This observation represents the discovery of a new spontaneous order phenomenon in soap film systems and provides a new energy-free approach for particle separation and soft colloidal crystal assembly.

High throughput clogging free microfluidic particle filter by femtosecond laser micromachining.

In recent decades, driven by the needs of industry and medicine, researchers have been investigating how to remove carefully from the main flow microscopic particles or clusters of them. Among all the approaches proposed, crossflow filtration is one of the most attractive as it provides a non-destructive, label-free and in-flow sorting method. In general, the separation performance shows capture and separation efficiencies ranging from 70% up to 100%. However, the maximum flow rate achievable (µL/min) is still orders of magnitude away from those suitable for clinical or industrial applications mainly due to the low stiffness of the materials typically used. In this work, we propose an innovative hydrodynamic-crossflow hybrid filter geometry, buried in a fused silica substrate by means of the femtosecond laser irradiation followed by chemical etching technique. The material high stiffness combined with the accuracy of our manufacturing technique allows the 3D fabrication of non-deformable channels with higher aspect ratio posts, while keeping the overall device dimensions compact. The filter performance has been validated through experiments with both Newtonian (water-based solution of microbeads) and non-Newtonian fluids (blood), achieving separation efficiencies of up to 94% and large particles recovery rates of 100%, even at very high flow rates (mL/h).

Laser microdissection pressure catapulting (LMPC): a new technique to handle single microplastic particles for number-based validation strategies.

This study examines laser microdissection pressure catapulting (LMPC) as an innovative method for microplastic research. Laser pressure catapulting as part of commercially available LMPC microscopes enables the precise handling of microplastic particles without any mechanical contact. In fact, individual particles with sizes between several micrometers and several hundred micrometers can be transported over centimeter-wide distances into a collection vial. Therefore, the technology enables the exact handling of defined numbers of small microplastics (or even individual ones) with the greatest precision. Herewith, it allows the production of particle number-based spike suspensions for method validation. Proof-of-principle LMPC experiments with polyethylene and polyethylene terephthalate model particles in the size range from 20 to 63 µm and polystyrene microspheres (10 µm diameter) demonstrated precise particle handling without fragmentation. Furthermore, the ablated particles showed no evidence of chemical alteration as seen in the particles' IR spectra acquired via laser direct infrared analysis. We propose LMPC as a promising new tool to produce future microplastic reference materials such as particle-number spiked suspensions, since LMPC circumvents the uncertainties resulting from the potentially heterogeneous behavior or inappropriate sampling from microplastic suspensions. Furthermore, LMPC could be advantageous for the generation of very accurate calibration series of spherical particles for microplastic analysis via pyrolysis-gas chromatography-mass spectrometry (down to 0.54 ng), as it omits the dissolution of bulk polymers.

Numerical Study of Particle Separation through Integrated Multi-Stage Surface Acoustic Waves and Modulated Driving Signals.

The manipulation of biomedical particles, such as separating circulating tumor cells from blood, based on standing surface acoustic wave (SSAW) has been widely used due to its advantages of label-free approaches and good biocompatibility. However, most of the existing SSAW-based separation technologies are dedicated to isolate bioparticles in only two different sizes. It is still challenging to fractionate various particles in more than two different sizes with high efficiency and accuracy. In this work, to tackle the problems of low efficiency for multiple cell particle separation, integrated multi-stage SSAW devices with different wavelengths driven by modulated signals were designed and studied. A three-dimensional microfluidic device model was proposed and analyzed using the finite element method (FEM). In addition, the effect of the slanted angle, acoustic pressure, and the resonant frequency of the SAW device on the particle separation were systemically studied. From the theoretical results, the separation efficiency of three different size particles based on the multi-stage SSAW devices reached 99%, which was significantly improved compared with conventional single-stage SSAW devices.

Phagosome proteomics to study Leishmania's intracellular niche in macrophages.

Intracellular pathogens invade their host cells and replicate within specialized compartments. In turn, the host cell initiates a defensive response trying to kill the invasive agent. As a consequence, intracellular lifestyle implies morphological and physiological changes in both pathogen and host cell. Leishmania spp. are medically important intracellular protozoan parasites that are internalized by professional phagocytes such as macrophages, and reside within the parasitophorous vacuole inhibiting their microbicidal activity. Whereas the proteome of the extracellular promastigote form and the intracellular amastigote form have been extensively studied, the constituents of Leishmania's intracellular niche, an endolysosomal compartment, are not fully deciphered. In this review we discuss protocols to purify such compartments by means of an illustrating example to highlight generally relevant considerations and innovative aspects that allow purification of not only the intracellular parasites but also the phagosomes that harbor them and analyze the latter by gel free proteomics.

Kinetics of solutes and particles of different size in the digestive tract of cattle of 0.5-10 years of age, and relationships with methane production.

The digestive physiology of cattle is characterised by comparatively long digesta mean retention times (MRTs), a particle sorting mechanism (difference in MRTs of large vs. small particles) and a distinct digesta washing (difference in MRTs between particles and fluids) in the reticulorumen (RR). How these processes mature during ontogeny, and how they link to other digestion characteristics and methane production, is largely unknown. We used a set of passage markers (Co-EDTA for fluids and hay particles of 2, 5 and 8 mm length mordanted with Cr, La and Ce, respectively) to measure MRTs in 12 heifers (0.5-2.1 years; hay only) and two groups of 15 lactating cows (2.4-10.0 years; forage-only vs. forage-concentrate diet). The MRTs differed between markers (Co < Cr < La < Ce) and were longer in heifers than cows, consistent with the lower feed intake in heifers. MRTs were mostly similar between cow groups and increased with age. Digesta washing was not affected by group, age, feed intake and number of chews per unit of feed. The degree of digesta washing was not related to CH4 measures. Particle sorting was more prominent in cows than heifers but did not differ between cow groups or change with age in cows. This could be the consequence of the abrupt increase in intake from heifers to cows at a time when gut capacity is not yet fully developed; particle sorting might then clear smaller particles from the RR sooner allowing a higher intake. Surprisingly, CH4 yield per ingested feed did not correlate with MRTs, and CH4 yield per unit of digested fibre decreased with increasing MRTs and with increasing fibre digestibility. As this pattern occurred in heifers and both cow groups, it appeared independent of age, indicating a mechanism that has not been described in the literature so far and requires further investigation.

Dielectrophoretic microbead sorting using modular electrode design and capillary-driven microfluidics.

Multiplexing assays using microbeads in microfluidics offers high flexibility and throughput, but requires the ability to sort particles based on their physical properties. In this paper, we present a continuous method for separating microbeads that is compact, modular and adaptive, employing an optimized electrode layout that alternates sorting and concentration of microbeads using dielectrophoresis and a nested design. By simulating the combined effects of the hydrodynamic drag and dielectrophoresis forces on polystyrene beads, the parameters of the electrode layout and voltage configuration are optimized for maximum separation based on particle size with a small number of slanted planar electrodes. Experimental verification confirms the efficient separation of 10 µm and 5 µm beads, with ~98% of all concentrated beads sorted in two separate streams and only ~2% of 5 µm beads leaking into the 10 µm bead stream. In addition, this method is implemented on capillary-driven microfluidic chips for maximum portability and ease of use.

A Radial Pillar Device (RAPID) for continuous and high-throughput separation of multi-sized particles.

Pillar-based microfluidic sorting devices are preferred for isolation of rare cells due to their simple designs and passive operation. Dead-end pillar filters can efficiently capture large rare cells, such as, circulating tumor cells (CTCs), nucleated red blood cells (NRBCs), CD4 cells in HIV patients, etc., but they get clogged easily. Cross flow filters are preferred for smaller rare particles (e.g. separating bacteria from blood), but they need additional buffer inlets and a large device footprint for efficient operation. We have designed a new microparticle separation device i.e. Ra dial Pi llar D evice (RAPID) that combines the advantages of dead-end and cross flow filters. RAPID can simultaneously isolate both large and small rare particles from a mixed population, while functioning for several hours without clogging. We have achieved simultaneous separation of 10 µ m and 2 µ m polystyrene particles from a mixture of 2 µ m, 7 µ m and 10 µ m particles. RAPID achieved average separation purity and recovery in excess of ∼90%. The throughput of our device (∼3ml/min) is 10 and 100 times higher compared to cross flow and dead-end filters respectively, thereby justifying the name RAPID.

A hybrid dielectrophoretic and hydrophoretic microchip for particle sorting using integrated prefocusing and sorting steps.

This work explores dielectrophoresis (DEP)-active hydrophoresis in sorting particles and cells. The device consists of prefocusing region and sorting region with great potential to be integrated into advanced lab-on-a-chip bioanalysis devices. Particles or cells can be focused in the prefocusing region and then sorted in the sorting region. The DEP-active hydrophoretic sorting is not only based on size but also on dielectric properties of the particles or cells of interest without any labelling. A mixture of 3 and 10 µm particles were sorted and collected from corresponding outlets with high separation efficiency. According to the different dielectric properties of viable and nonviable Chinese Hamster Ovary (CHO) cells at the medium conductivity of 0.03 S/m, the viable CHO cells were focused well and sorted from cell sample with a high purity.

Isolation of Phytochrome B Photobodies.

Phytochrome B (phyB), a plant photoreceptor, forms a membraneless organelle known as a photobody. Here, we present a protocol for the isolation of phyB photobodies through fluorescence-activated particle sorting from mature transgenic Arabidopsis leaves expressing phyB-GFP. This protocol involves the isolation of nuclei from frozen ground leaves using sucrose gradient centrifugation, the disruption of nuclear envelopes by sonication, and the subsequent isolation of phyB photobodies through fluorescence-activated particle sorting. We include experimental tips and notes for each step.

Vault-phagy: a phase-separation-mediated selective autophagy of vault, a non-membranous organelle.

SQSTM1/p62 bodies are phase-separated condensates that play a fundamental role in intracellular quality control and stress responses. Despite extensive studies investigating the mechanism of formation and degradation of SQSTM1/p62 bodies, the constituents of SQSTM1/p62 bodies remain elusive. We recently developed a purification method for intracellular SQSTM1/p62 bodies using a cell sorter and identified their constituents by mass spectrometry. Combined with mass spectrometry of tissues from selective autophagy-deficient mice, we identified vault, a ubiquitous non-membranous organelle composed of proteins and non-coding RNA, as a novel substrate for selective autophagy. Vault directly binds to NBR1, an SQSTM1/p62 binding partner recruited to SQSTM1/p62 bodies, and is subsequently degraded by selective autophagy dependent on the phase separation of SQSTM1/p62. We named this process "vault-phagy" and found that defects in vault-phagy are related to nonalcoholic steatohepatitis (NASH)-derived hepatocellular carcinoma. Our method for purifying SQSTM1/p62 bodies will contribute to elucidating the mechanisms of several stress responses and diseases mediated by SQSTM1/p62 bodies.

Response of soil particles around bedrock outcrops to sorting of rock surface flow derived outcrops in a rocky desertification area.

Soils around bedrock outcrops, even if they are protected by vegetation to some extent after ecological restoration, are prone to being washed away by rock surface flow (RSF) derived from these outcrops in rocky desertification land. However, the extent of the scouring scale and sorting effect of RSF on the soils around outcrops remains unknown. To solve this problem, a series of soils around bedrock outcrops exposed in sloping farmland (SF, without RSF), abandoned land (AL, 1 year of RSF) and shrub-grassland (SG, 5 years of RSF) were examined by the laser diffraction method in a natural ecological restoration area of rocky desertification, where the duration of the RSF is also the time for ecological restoration. It was found that the RSF had a limited effect on the particle size distribution of the soils, only having a significant scouring effect on the soils at the rock-soil interface within a horizontal distance of 2 cm from the outcrops and an insignificant effect on the soils far away from the outcrops in terms of horizontal distance (10 cm and 20 cm). The particle size distributions of the soil around the outcrops were related to erosion caused by the RSF, but mainly benefited from ecological restoration. Compared with SF, the fine particle content in the soils around the outcrops significantly decreased in AL, but significantly increased in SG. Within a short period (1 year) after natural recovery, the RSF had a reduced effect on the fine particles of the soil around the outcrops; however, this did not occur after a long period (5 years). The results of this study further explain the influence of the RSF on soil erosion and leakage loss in karst areas.

Method to Purify Phase-Separated p62 Bodies Using Fluorescence-Activated Particle Sorting.

p62 bodies are ubiquitin-positive cytoplasmic condensates formed by liquid-liquid phase separation. They are targeted by selective autophagy and play important roles in intracellular quality control and stress responses. However, little is known about their constituents. In this chapter, we describe a method for purifying p62 bodies using fluorescence-activated particle sorting. This method contributes to the identification of novel components of p62 bodies under various physiological and stress conditions.

Particle sorting to improve the removal of fluoride and aluminum nitride from secondary aluminum dross by roasting.

It is important to remove active substances from secondary aluminum dross (SAD) to meet the reuse of SAD. In this work, the removal of active substances from different particle sizes of SAD was studied using roasting improvement with particle sorting. The results showed that roasting after particle sorting pretreatment can effectively remove fluoride and aluminum nitride (AlN) from SAD, while getting the high-grade alumina (Al2O3) crude materials. The active substances of SAD mainly contribute to AlN, aluminum carbide (Al4C3), and soluble fluoride ions. AlN and Al3C4 mainly exist in particles of 0.05-0.1 mm, while Al and fluoride are mainly in particles of 0.1-0.2 mm. The SAD of particle size ranging 0.1-0.2 mm has high activity and leaching toxicity; the gas emission was reached 50.9 mL/g (limit value of 4 mL/g), and the fluoride ion concentration in the literature was 137.62 mg/L (limit value of 100 mg/L) during the identification for reactivity and leaching toxicity according to GB5085.5-2007 and GB5085.3-2007, respectively. Roasting at 1000 °C for 90 min, the active substances of SAD were converted to Al2O3, N2, and CO2; meanwhile, soluble fluoride converted to stable CaF2. The final gas release was reduced to 2.01 mL/g while soluble fluoride from SAD residues was reduced to 6.16 mg/L, respectively. The Al2O3 content of SAD residues was determined at 91.8% and has been classified as category I solid waste. The results suggested that the roasting improvement with particle sorting of SAD can meet the reuse of valuable materials at full scale.

The Physics and Manipulation of Dean Vortices in Single- and Two-Phase Flow in Curved Microchannels: A Review.

Microchannels with curved geometries have been employed for many applications in microfluidic devices in the past decades. The Dean vortices generated in such geometries have been manipulated using different methods to enhance the performance of devices in applications such as mixing, droplet sorting, and particle/cell separation. Understanding the effect of the manipulation method on the Dean vortices in different geometries can provide crucial information to be employed in designing high-efficiency microfluidic devices. In this review, the physics of Dean vortices and the affecting parameters are summarized. Various Dean number calculation methods are collected and represented to minimize the misinterpretation of published information due to the lack of a unified defining formula for the Dean dimensionless number. Consequently, all Dean number values reported in the references are recalculated to the most common method to facilitate comprehension of the phenomena. Based on the converted information gathered from previous numerical and experimental studies, it is concluded that the length of the channel and the channel pathline, e.g., spiral, serpentine, or helix, also affect the flow state. This review also provides a detailed summery on the effect of other geometric parameters, such as cross-section shape, aspect ratio, and radius of curvature, on the Dean vortices' number and arrangement. Finally, considering the importance of droplet microfluidics, the effect of curved geometry on the shape, trajectory, and internal flow organization of the droplets passing through a curved channel has been reviewed.

Chiral Mechanical Effect of the Tightly Focused Chiral Vector Vortex Fields Interacting with Particles.

The coupling of the spin-orbit angular momentum of photons in a focused spatial region can enhance the localized optical field's chirality. In this paper, a scheme for producing a superchiral optical field in a 4π microscopic system is presented by tightly focusing two counter-propagating spiral wavefronts. We calculate the optical forces and torques exerted on a chiral dipole by the chiral light field and reveal the chiral forces by combining the light field and dipoles. Results indicate that, in addition to the general optical force, particles' motion would be affected by a chiral force that is directly related to the particle chirality. This chiral mechanical effect experienced by the electromagnetic dipoles excited on a chiral particle could be characterized by the behaviors of chirality density and flux, which are, respectively, associated with the reactive and dissipative components of the chiral forces. This work facilitates the advancement of optical separation and manipulation techniques for chiral particles.

Low-frequency electrokinetics in a periodic pillar array for particle separation.

Deterministic Lateral Displacement (DLD) exploits periodic arrays of pillars inside microfluidic channels for high-precision sorting of micro- and nano-particles. Previously we demonstrated how DLD separation can be significantly improved by the addition of AC electrokinetic forces, increasing the tunability of the technique and expanding the range of applications. At high frequencies of the electric field (>1 kHz) the behaviour of such systems is dominated by Dielectrophoresis (DEP), whereas at low frequencies the particle behaviour is much richer and more complex. In this article, we present a detailed numerical analysis of the mechanisms governing particle motion in a DLD micropillar array in the presence of a low-frequency AC electric field. We show how a combination of Electrophoresis (EP) and Concentration-Polarisation Electroosmosis (CPEO) driven wall-particle repulsion account for the observed experimental behaviour of particles, and demonstrate how this complete model can predict conditions that lead to electrically induced deviation of particles much smaller than the critical size of the DLD array.

Separation of bacteria smaller than 4 µm from other blood components using insulator-based dielectrophoresis: numerical simulation approach.

Bloodstream infection (BSI) is a life-threatening infection that causes more than 80,000 deaths and more than 500,000 infections annually in North America. The rapid diagnosis of infection reduces BSI mortality. We proposed bacterial enrichment and separation approach in the current work that may reduce culturing time and accelerate the diagnosis of infection. Over the last two decades, multiple separation methods have been developed, and among these methods, insulator-based dielectrophoresis (iDEP) is considered a powerful technique for separating biological particles. Bacterial separation in the blood is challenging due to the presence of other blood cells, such as white blood cells, red blood cells, and platelets. In the present study, a model is presented which is capable of blood cells separation and directing each cell to a specific outlet using continuous flows of particles with sizes larger than 8 µm, 8-4 µm, and smaller than 4 µm. Compared to other methods, such as filtration, the main advantage of this model is that particles larger than 8 µm are separated from the flow before other particles, which prevents the accumulation of particles in the channel. The outcomes of simulations demonstrated that the factors such as applied voltage and channel dimensions significantly affect the separation efficiency. If these values are properly selected (for example voltage of 70 V that was causing an electric field of 200 V/cm), the proposed model can completely (100%) separate particles larger than 8 µm and smaller than 4 µm (8-4 µm particles separation efficiency is 95%).

Integrated proteomics identifies p62-dependent selective autophagy of the supramolecular vault complex.

In addition to membranous organelles, autophagy selectively degrades biomolecular condensates, in particular p62/SQSTM1 bodies, to prevent diseases including cancer. Evidence is growing regarding the mechanisms by which autophagy degrades p62 bodies, but little is known about their constituents. Here, we established a fluorescence-activated-particle-sorting-based purification method for p62 bodies using human cell lines and determined their constituents by mass spectrometry. Combined with mass spectrometry of selective-autophagy-defective mouse tissues, we identified vault, a large supramolecular complex, as a cargo within p62 bodies. Mechanistically, major vault protein directly interacts with NBR1, a p62-interacting protein, to recruit vault into p62 bodies for efficient degradation. This process, named vault-phagy, regulates homeostatic vault levels in vivo, and its impairment may be associated with non-alcoholic-steatohepatitis-derived hepatocellular carcinoma. Our study provides an approach to identifying phase-separation-mediated selective autophagy cargoes, expanding our understanding of the role of phase separation in proteostasis.

Continuous Particle Separation Driven by 3D Ag-PDMS Electrodes with Dielectric Electrophoretic Force Coupled with Inertia Force.

Cell separation has become @important in biological and medical applications. Dielectrophoresis (DEP) is widely used due to the advantages it offers, such as the lack of a requirement for biological markers and the fact that it involves no damage to cells or particles. This study aimed to report a novel approach combining 3D sidewall electrodes and contraction/expansion (CEA) structures to separate three kinds of particles with different sizes or dielectric properties continuously. The separation was achieved through the interaction between electrophoretic forces and inertia forces. The CEA channel was capable of sorting particles with different sizes due to inertial forces, and also enhanced the nonuniformity of the electric field. The 3D electrodes generated a non-uniform electric field at the same height as the channels, which increased the action range of the DEP force. Finite element simulations using the commercial software, COMSOL Multiphysics 5.4, were performed to determine the flow field distributions, electric field distributions, and particle trajectories. The separation experiments were assessed by separating 4 µm polystyrene (PS) particles from 20 µm PS particles at different flow rates by experiencing positive and negative DEP. Subsequently, the sorting performances of the 4 µm PS particles, 20 µm PS particles, and 4 µm silica particles with different solution conductivities were observed. Both the numerical simulations and the practical particle separation displayed high separating efficiency (separation of 4 µm PS particles, 94.2%; separation of 20 µm PS particles, 92.1%; separation of 4 µm Silica particles, 95.3%). The proposed approach is expected to open a new approach to cell sorting and separating.