Suspension Conditions Research Articles

Dextran sulfate prevents excess aggregation of human pluripotent stem cells in 3D culture by inhibiting ICAM1 expression coupled with down-regulating E-cadherin through activating the Wnt signaling pathway

BackgroundHuman pluripotent stem cells (hPSCs) have great potential in applications for regenerative medicine and drug development. However, 3D suspension culture systems for clinical-grade hPSC large-scale production have been a major challenge. Accumulating evidence has demonstrated that the addition of dextran sulfate (DS) could prevent excessive adhesion of hPSCs from forming larger aggregates in 3D suspension culture. However, the signaling and molecular mechanisms underlying this phenomenon remain elusive.MethodsBy using a cell aggregate culture assay and separating big and small aggregates in suspension culture systems, the potential mechanism and downstream target genes of DS were investigated by mRNA sequence analysis, qRT-PCR validation, colony formation assay, and interference assay.ResultsSince cellular adhesion molecules (CAMs) play important roles in hPSC adhesion and aggregation, we assumed that DS might prevent excess adhesion through affecting the expression of CAMs in hPSCs. As expected, after DS treatment, we found that the expression of CAMs was significantly down-regulated, especially E-cadherin (E-cad) and intercellular adhesion molecule 1 (ICAM1), two highly expressed CAMs in hPSCs. The role of E-cad in the adhesion of hPSCs has been widely investigated, but the function of ICAM1 in hPSCs is hardly understood. In the present study, we demonstrated that ICAM1 exhibited the capacity to promote the adhesion in hPSCs, and this adhesion was suppressed by the treatment with DS. Furthermore, transcriptomic analysis of RNA-seq revealed that DS treatment up-regulated genes related to Wnt signaling resulting in the activation of Wnt signaling in which SLUG, TWIST, and MMP3/7 were highly expressed, and further inhibited the expression of E-cad.ConclusionOur results demonstrated that DS played an important role in controlling the size of hPSC aggregates in 3D suspension culture by inhibiting the expression of ICAM1 coupled with the down-regulation of E-cad through the activation of the Wnt signaling pathway. These results represent a significant step toward developing the expansion of hPSCs under 3D suspension condition in large-scale cultures.

Transition from serum-supplemented monolayer to serum-free suspension lentiviral vector production for generation of chimeric antigen receptor T cells

Background aimsLentiviral vectors (LVs) have been used extensively in gene therapy protocols because of their high biosafety profile and capacity to stably express a gene of interest. Production of these vectors for the generation of chimeric antigen receptor (CAR) T cells in academic and research centers is achieved using serum-supplemented static monolayer cultures. Although efficient for pre-clinical studies, this method has a number of limitations. The main hurdles are related to its incompatibility with robust and controlled large-scale production. For this reason, cell suspension culture in bioreactors is desirable. Here the authors report the transition of LV particle production from serum-supplemented monolayer to serum-free suspension culture with the objective of generating CAR T cells. MethodsA self-inactivating LV anti-CD19 CAR was produced by transient transfection using polyethylenimine (PEI) in human embryonic kidney 293 T cells previously adapted to serum-free suspension culture. ResultsLV production of 8 × 106 transducing units (TUs)/mL was obtained in serum-supplemented monolayer culture. LV production in the serum-free suspension conditions was significantly decreased compared with monolayer production. Therefore, optimization of the transfection protocol was performed using design of experiments. The results indicated that the best condition involved the use of 1 μg of DNA/106 cells, 1 × 106 cells/mL and PEI:DNA ratio of 2.5:1. This condition used less DNA and PEI compared with the standard, thereby reducing production costs. This protocol was further improved with the addition of 5 mM of sodium butyrate and resulted in an increase in production, with an average of 1.5 × 105 TUs/mL. LV particle functionality was also assessed, and the results indicated that in both conditions the LV was capable of inducing CAR expression and anti-tumor response in T cells, which in turn were able to identify and kill CD19+ cells in vitro. ConclusionsThis study demonstrates that the transition of LV production from small-scale monolayer culture to scalable and controllable bioreactors can be quite challenging and requires extensive work to obtain satisfactory production.

Microprobe‐based technology for antigen‐specific purification of exosomes for OMICS analysis

Exosomes encapsulate genomic and proteomic biomarkers for non‐invasive diagnosis and disease monitoring. However, exosome surface‐markers heterogeneity is a major drawback of current isolation methods. Here, we report a direct, one‐step exosome sampling technology, ExoPRIME, for selective capture of CD63+ exosome subpopulations using an immune‐affinity protocol. Microneedles (300μm × 30 mm), functionalized with anti‐ CD63 antibodies, were incubated under various experimental conditions in conditioned astrocyte medium and astrocyte‐derived exosome suspension. The probe’s capture efficiency and specificity were validated using Flu‐oroCet assay, immunofluorescent imaging, and OMICS analyses. Significantly higher exosomes were captured by probes incubated for 16 h at 4 0C in enriched exosomal suspension (23 × 10 6 exosomes per probe) vis‐'a‐vis 2 h at 4 0 C (12 × 10 6) and 16 h at 22 0C (3 × 10 6) in conditioned cell media. Our results demonstrate the application of ExoPRIME over a broad dynamic range of temperature and incubation parameters, offering flexibility for any desired application. ExoPRIME permits the use and re‐use of minimal sample volumes (≤200 μL), can be multiplexed in arrays, and integrated into a lab‐on‐a‐chip platform to achieve parallel, high‐throughput isolation of different exosome classes in a semi‐automated workstation. This platform could provide direct exosomal analysis of biological fluids since it can elegantly interface with existing room‐temperature, picomolar‐range nucleic acid assays to provide a clinical diagnostic tool at the point of care.

Conditions for suspension of forced execution Procedure

The suspension of enforcement proceedings means temporarily halting enforcement actions until the grounds for suspension laid down by law and indicated in the document of suspension have ceased. The jurisdiction to suspend the enforcement proceedings lies with the bailiff under Article 78 EC (mandatory suspension) and with the court for the grounds laid down in Article 79 EC (facultative suspension). The appearance of the circumstances listed by law establishes the unconditional obligation of the bailiff to suspend the enforcement procedure. The question of suspension is dealt with by the bailiff of his own initiative or on request, by notifying the creditor and the debtor. The facultative suspension of enforcement proceedings is a right of the court. The decision of granting the request for suspension of enforcement proceedings prevents the enforcement proceedings from taking place until the circumstances for which it was issued have ceased to exist.

Efficiency of Financing Transactions of the Real Sector of the Economy with the Use of a Currency Barrier Option in the Conditions of Economic Imbalance 2013—2018

Goals. A comprehensive author’s study of the eff ectiveness of financing transactions in the real sector of the economy using a currency option with a barrier suspensive condition in conditions of economic imbalance. Assessment of the Russian practice of insuring market risks throughover-the-counter derivative financial instruments. Methods. The study used a mathematical model of Rayner and Rubinstein were used to calculate the fair value of options. Results. The sale of a call option of the «foreign exchange delivery option with a barrier condition» type allows the company to receive an option premium, but at the same time the company assumes an unlimited risk of losses. If the transaction is concluded by the parties for speculative purposes, then the symmetry and balance of the positions is achieved due to the amount of the premium paid by the buyer. The recommendations given in the article on the use of currency barrier options and the assessment of their value will improve the efficiency of market risk management systems in real sector companies. Conclusions. In the period from 2013 to 2018, companies actively used derivative financial instruments in their practice, both for the purpose of hedging or obtaining an option premium, and for the purpose of reducing the cost of ruble financing. Incorrect assessment of their risks, insufficient experience in the use and evaluation of derivatives led to the fact that by the end of 2014—2018, in fact, most of the largest Russian companies suff ered a loss from the use of derivative financial instruments. The competent use of derivatives for the purposes of hedging market risks will allow companies toavoid losses in conditions of increased risk.

Mesogels control volume to explore the glass transition

Suspending colloidal particles in solution with thermosensitive hydrogel spheres allows for study of the liquid and solid conditions in glass-forming suspensions

Novel microgel culture system as semi-solid three-dimensional in vitro model for the study of multiple myeloma proliferation and drug resistance.

Multiple myeloma (MM) is a hematological malignancy in which the patient's drug resistance is one of the main clinical problems. As 2D cultures do not recapitulate the cellular microenvironment, which has a key role in drug resistance, there is an urgent need for better biomimetic models. Here, a novel 3D platform is used to model MM. The semi-solid culture consists of a dynamic suspension of microspheres and MM cells, termed as microgel. Microspheres are synthesized with acrylic polymers of different sizes, compositions, and functionalities (fibronectin or hyaluronic acid). Optimal conditions for the platform in terms of agitation speed and microsphere size have been determined. With these parameters the system allows good proliferation of the MM cell lines RPMI8226, U226, and MM1.S. Interestingly, when used for drug resistance studies, culture of the three MM cell lines in microgels showed close agreement in revealing the role of acrylic acid in resistance to anti-MM drugs such as dexamethasone and bortezomib. This work presents a unique platform for the in vitro modeling of non-solid tumors since it allows keeping non-adherent cells in suspension conditions but in a 3D context that can be easily tuned with different functionalizations.

Technology of primary water purification

The polluted wastewater treatment is one of the main issues in agro-industrial complex. This article is devoted to the primary water purification research for its further treatment with ferrate technology. The technology implies the wastewater treatment from household and industrial waste resulting from agricultural enterprises. The ferrate technology provides a high efficiency of pretreated water purification, in comparison with other existing methods. The article substantiates such an advantage accordingly. The waste generated as a result of wastewater treatment is non-toxic and disposal. The article proposes a new method of preliminary water purification based on its treatment between a galvanic couple in the presence of the calcium hydroxide. This method can be used at water treatment plants and at enterprises carrying out industrial activities in agriculture. The method is applicable only after mechanical filtration at the stage of primary water treatment. The proposed method provides water treatment from a wide range of various origins contaminants. The article also describes an experimental equipment specially designed for the proposed method’s implementation and further research. The equipment provides continuous water treatment consuming electricity, quicklime and compressed air. To maintain the lime particles in suspension condition the airflow is supplied through an aerator. The article applies the turbidimetric analysis method to conduct an in-depth analysis of the water treatment processes and determine the relative reagent concentration in the working area. The article also proves the absence of interface formation between water and milk of lime when subjected to air flow.

Preparation of Natural Rubber Composites with High Silica Contents Using a Wet Mixing Process.

A wet mixing process is proposed for filled rubber composites with a high silica loading to overcome the drawbacks of high energy consumption and workplace contamination of the conventional dry mixing process. Ball milling was adopted for preparing the silica dispersion because it has a simple structure, is easy to operate, and is a low-cost process that can be easily scaled up for industrial production. The response surface methodology was used to optimize the making of the silica dispersion. The optimum conditions for a well-dispersed silica suspension with the smallest silica particle size of 4.9 mm were an about 22% silica content and 62 h of ball milling. The effects of dry and wet mixing methods on the properties of silica-filled rubber composites were investigated in a broad range of silica levels from low to high loadings. The mixing method choice had little impact on the properties of rubber composites with low silica loadings. The silica-filled rubber demonstrated in this study, however, shows superior characteristics over the rubber composite prepared with conventional dry mixing, particularly with high silica loadings. When compared to silica-filled natural rubbers prepared by dry mixing (dry silica rubber, DSR), the wet mixing (for WSR) produced smaller silica aggregates with better dispersion. Due to the shorter heat history, the WSR exhibits superior curing characteristics such as a longer scorch time (2.2–3.3 min for WSR and 1.0–2.1 min for DSR) and curing time (4.1–4.5 min for WSR and 2.2–3.1 min for DSR). Additionally, the WSR has superior mechanical properties (hardness, modulus, tensile strength, and especially the elongation at break (420–680% for WSR and 360–620% DSR)) over the DSR. The rolling resistance of WSR is lower than that of DSR. However, the reversed trend on the wet skid resistance is observed.

CAN-Based Vibration Control for Networked Vehicle Active Suspension with Both Network-Induced Delays and Packet-Dropouts

Ride comfort and driving safety are highly vulnerable to the undesirable excessive vibrations caused by road surface irregularities and the imperfect in-vehicle network (IVN). The main contribution of this paper consists of proposing a near-optimal vibration control approach for networked vehicle active suspension under irregular road excitations in a discrete-time domain, in which the uncertain time delay and packet dropout in CAN are taken into consideration. More specially, by virtue of two buffers of the sensor-to-controller network channel and the controller-to-actuator network channel in CAN, by introducing a designed state-transformation-based method, the original vibration control problem under the constraints of the irregular road excitations and imperfect CAN is transformed into a two-point boundary value (TPBV) problem without advanced and delayed items. After that, the near-optimal vibration control approach is presented to isolate the vehicle body from the road excitations and compensate the time delay and packet dropout from CAN synchronously. The stability condition of the networked vehicle active suspension under the proposed vibration controller is obtained based on the Lyapunov function. In numerous scenarios with different road roughnesses and network-induced time delays and packet dropouts, the simulation results illustrate the effectiveness and superiority of the proposed near-optimal vibration controller.

The electrostatic levitation system for active suspension control of test sample in horizontal and vertical axes

An electrostatic levitation system (ESLS) with active control model is proposed to realize the noncontact suspension of test sample for analyzing material properties. The characteristics about the electric field force and the electric image force of ESLS are firstly analyzed, and then the dynamic functions of test sample in the ESLS are developed. Furthermore, the conditions of test sample’s stable suspensions in horizontal and vertical directions are proposed, it shows that the control voltage is regulated to guarantee suspension stability of test sample considering the gravity and the displacement deflection in vertical direction. The control models of test sample in the ESLS are established to realize the active suspension in vertical and horizontal directions. Finally, both simulation and experiment with different suspension conditions are conducted to test the suspension characteristics of test sample in ESLS, and results indicate that the ESLS could stably suspend the test sample through regulating control voltages of high voltage electrodes and the displacements of test sample. Therefore, the ESLS could be used as a noncontact holder of test sample in materials testing instruments.

A Redox‐Active Microporous Organosiloxane Containing a Stable Neutral Radical, Trioxotriangulene

A new silyl-substituted trioxotriangulene (TOT) neutral radical and corresponding porous organosiloxanes (POSs) were synthesized. The neutral radical exhibited a peculiarly high stability and formed a diamagnetic π-dimer characteristic to TOT neutral radicals stabilized by the strong multiple SOMO-SOMO interaction in both solution and solid states. POSs including TOT units within the organosiloxane-wall were prepared by polycondensation of the silyl groups and formed microporous structures with ∼1 nm-size diameters. Redox ability of TOT units in the POS was demonstrated by the treatment of oxidant/reductant in heterogeneous suspension condition, where the TOT units were reversibly converted between reduced and neutral radical species. Furthermore, the solid-state electrochemical measurements of the POS revealed the reversible multi-stage redox ability of TOT units involving polyanionic species within the organosiloxane-wall.

Machine learning as a tool to engineer microstructures: Morphological prediction of tannin-based colloids using Bayesian surrogate models

Oxidized tannic acid (OTA) is a useful biomolecule with a strong tendency to form complexes with metals and proteins. In this study we open the possibility to further the application of OTA when assembled as supramolecular systems, which typically exhibit functions that correlate with shape and associated morphological features. We used machine learning (ML) to selectively engineer OTA into particles encompassing one-dimensional to three-dimensional constructs. We employed Bayesian regression to correlate colloidal suspension conditions (pH and pKa) with the size and shape of the assembled colloidal particles. Fewer than 20 experiments were found to be sufficient to build surrogate model landscapes of OTA morphology in the experimental design space, which were chemically interpretable and endowed predictive power on data. We produced multiple property landscapes from the experimental data, helping us to infer solutions that would satisfy, simultaneously, multiple design objectives. The balance between data efficiency and the depth of information delivered by ML approaches testify to their potential to engineer particles, opening new prospects in the emerging field of particle morphogenesis, impacting bioactivity, adhesion, interfacial stabilization, and other functions inherent to OTA.Impact statementTannic acid is a versatile bio-derived material employed in coatings, surface modifiers, and emulsion and growth stabilizers, which also imparts mild anti-viral health benefits. Our recent work on the crystallization of oxidized tannic acid (OTA) colloids opens the route toward further valuable applications, but here the functional properties tend to depend strongly on particle morphology. In this study, we eschew trial-and-error morphology exploration of OTA particles in favor of a data-driven approach. We digitalized the experimental observations and input them into a Gaussian process regression algorithm to generate morphology surrogate models. These help us to visualize particle morphology in the design space of material processing conditions, and thus determine how to selectively engineer one-dimensional or three-dimensional particles with targeted functionalities. We extend this approach to visualize other experimental outcomes, including particle yield and particle surface-to-volume ratio, which are useful for the design of products based on OTA particles. Our findings demonstrate the use of data-efficient surrogate models for general materials engineering purposes and facilitate the development of next-generation OTA-based applications.Graphic abstract

Circulating Tumor Cell Lines: an Innovative Tool for Fundamental and Translational Research.

Metastasis is a leading cause of cancer death. Despite improvements in treatment strategies, metastatic cancer has a poor prognosis. We thus face an urgent need to understand the mechanisms behind metastasis development, and thus to propose efficient treatments for advanced cancer. Metastatic cancers are hard to treat, as biopsies are invasive and inaccessible. Recently, there has been considerable interest in liquid biopsies including both cell-free circulating deoxyribonucleic acid (DNA) and circulating tumor cells from peripheral blood and we have established several circulating tumor cell lines from metastatic colorectal cancer patients to participate in their characterization. Indeed, to functionally characterize these rare and poorly described cells, the crucial step is to expand them. Once established, circulating tumor cell (CTC) lines can then be cultured in suspension or adherent conditions. At the molecular level, CTC lines can be further used to assess the expression of specific markers of interest (such as differentiation, epithelial or cancer stem cells) by immunofluorescence or cytometry analysis. In addition, CTC lines can be used to assess drug sensitivity to gold-standard chemotherapies as well as to targeted therapies. The ability of CTC lines to initiate tumors can also be tested by subcutaneous injection of CTCs in immunodeficient mice. Finally, it is possible to test the role of specific genes of interest that might be involved in cancer dissemination by editing CTC genes, by short hairpin ribonucleic acid (shRNA) or Crispr/Cas9. Modified CTCs can thus be injected into immunodeficient mouse spleens, to experimentally mimic part of the metastatic development process in vivo. In conclusion, CTC lines are a precious tool for future research and for personalized medicine, where they will allow prediction of treatment efficiency using the very cells that are originally responsible for metastasis.

Effect of pipeline surface roughness on peak impact forces caused by hydrodynamic submarine mudflow

The effect of pipeline surface roughness on the interaction between submarine landslides and pipelines may not be insignificant and has rarely been quantified. In this paper, a previously validated computational fluid dynamics model is employed to study the interaction of submarine landslides and suspended or laid-on-seabed pipelines, respectively, which are roughened in order to quantify the effect of pipeline surface roughness. Here, four orders of magnitude surface roughnesses are chosen whereas the mudflows (submarine landslides) occur at typical Reynolds numbers. The effect of surface roughness is primarily reflected in the peak load of the impact forces on the pipelines, which are more sensitive to high Reynolds numbers and suspension conditions. For suspended pipelines, with the increase of roughness, (i) the peak lift force increases, (ii) the peak drag force decreases, and (iii) Strouhal number slightly increases. Compared with nearly smooth pipelines, the relative increase of the peak lift force and the relative reduction of the peak drag force on suspended pipelines with a surface roughness of 0.15 mm attain 62% or 17%, respectively. Additionally, relevant mechanisms are highlighted via contours of the pressure around the pipeline, the interfacial shear rate, as well as the squeeze and hindrance effect of the seabed. Further, a standard chart methodology considering pipeline roughness to estimate peak impact forces is established, which provides a basis for the risk assessment of the whole life cycle of submarine pipelines.

The Donor’s Right to Reside for Life as a Condition in the Gift Agreement

The paper discusses the issue of the possibility of including the donor’s right to live for life in the real estate gift agreement. The relevance of this topic is predetermined by the large number of legal disputes that arise after the conclusion of a gift agreement in connection with the eviction of the donor from the living premises. There is no common point of view among theorists about the admissibility of the condition of life-long residence in the gift agreement, since this condition can be regarded as consideration. The situation is complicated by the fact that the right to live for life is not mentioned either in the Civil Code of the Russian Federation or in the Housing Code of the Russian Federation. Nevertheless, the courts, as a rule, recognize this condition in the contract as valid. It is proposed to include a canceling or suspensive condition in the gift agreement. Under the canceling condition, in case of violation of the donor’s right to live for life, the legal relationship terminates from the moment of violation of the right. The suspensive condition stipulates the following: in case of violation of the right of life-long residence, the donor has the right to cancel the gift. This will allow, firstly, to secure for the donor the right to cancel the gift if his right to live for life is violated. Secondly, this will resolve theoretical disputes, since the canceling and suspensive conditions are supplementary and do not change the causation of the transaction.

Experimental Study on the Suspension Amount of Invasion Gas in Drilling Fluid with Yield Stress

The gas suspension phenomenon caused by the yield stress of the drilling fluid affects the accurate calculation of wellbore pressure after gas invasion. At present, most studies on the bubble suspension in the yield stress fluid focus on the single-bubble suspension condition and there are few studies on the gas suspension concentration. This paper carried out the GSC (gas suspension concentration) experiment in the simulated drilling fluid, xanthan solution, with different gas invasion methods. The GSC in the drilling fluid under the conditions of diffuse gas invasion and differential pressure gas invasion was simulated by using two methods of stir-depressurization and continuous ventilation. The results showed that when the size of a single bubble satisfied the single-bubble suspension condition, multiple bubbles can be suspended at the same time. The GSC is affected by the average size of the suspended bubbles, the yield stress of the drilling fluid, and the gas invasion modes. For different gas invasion modes, the empirical models of critical GSC related to the dimensionless number Bi are established. Compared with the experimental data, the relative error of the critical GSC in diffuse gas invasion is less than 6% and the relative error of the critical GSC in differential pressure gas invasion is less than 10%. The results of this work can provide guiding significance for accurate calculation of wellbore pressure.

AUCTION SALES “SUBJECT TO CONFIRMATION” AND FORMATION OF CONTRACTS Withok Small Farms (Pty) Ltd v Amber Sunrise Properties 5 (Pty) Ltd 2009 2 SA 504 (SCA)

The Supreme Court of Appeal’s judgment in Withok Small Farms (Pty) Ltd v Amber Sunrise Properties 5 (Pty) Ltd (2009 2 SA 504 (SCA) (“Withok”)) raises two important issues for property practitioners, the one relating to auction sales and the other to the formation of contracts. The dispute arose largely because of a badly drafted agreement of sale document, but the import of the judgment is such that it may be prudent for practitioners to revisit even their well-drafted standard form sale and lease documents. The facts were straightforward. Certain properties owned by the first and second appellants (“the sellers”) were put up for sale at a public auction on 13 June 2006. The respondent, represented by one A, put in the highest bid, which the auctioneer accepted. Both A and the auctioneer signed a document entitled “Agreement and Conditions of Sale” (“the conditions of sale”), which set out the conditions relating to the auction. On 20 June 2006 the sellers’ representative confirmed the sale in writing by adding his signature in the allotted space on the final page of the document. However, the confirmation of the sale was not communicated to the respondent within the time contemplated in clause 1. In fact, the respondent did not receive notice of the confirmation until some time early in July 2006.The respondent did not wish to be bound by the sale and in due course applied for an order declaring the agreement to be of no force and effect. Its case was that the confirmation of the sale had not been communicated to it within the seven-day period contemplated in clause 1 of the conditions of sale, with the result that no agreement came into existence. The sellers, in turn, contended that the conditions of sale signed by the respondent and the auctioneer at the time of the auction constituted an agreement of salesubject to a suspensive condition, namely the confirmation of the sale by the sellers; the condition was fulfilled immediately upon the confirmation of the sale and without any need for it to be communicated to the respondent. The only issue in dispute was whether or not the confirmation of the sale had to be communicated to the respondent within the seven-day period.

Two-compartment membrane electrochemical remediation of heavy metals from an aged electroplating-contaminated soil: A comparative study of anodic and cathodic processes

In this study, two-compartment membrane electrochemical remediation (MER) based on the anode process and the cathode process strategies were compared for treating a multi metal -contaminated soil. Remediation effect, as well as energy consumption and risk evaluation of the two strategies under different current density conditions of electroplating-contaminated soil suspension were performed, the following conclusions were drawn. MERs based on both the anode and cathode processes exhibited a synergetic effect because the DC electric field and extractants dissolved more metals from the soil phase into the liquid phase of the suspension compared to a usual soil washing treatment. The maximum Cr, Cu, and Ni removal efficiencies of MERs based on the anode process were 79.5%, 86.2%, and 85.0%, respectively, compared to 27.5%, 72.5%, and 65.9% based on the cathode process. Risk assessment results showed lower soil environmental risk after MER based on the cathode process than after MER based on the anode process. In this study, MER based on the cathode process as an evolving soil remediation strategy was found to present high simultaneous remediation ability for soil heavy metals and leaching materials, showing its advantages of environmental friendliness and economic effectiveness.

ExoPRIME: Solid-phase immunoisolation and OMICS analysis of surface-marker-specific exosomal subpopulations

Exosomes encapsulate genomic and proteomic biomarkers for non-invasive diagnosis and disease monitoring. However, exosome surface-markers heterogeneity is a major drawback of current isolation methods. Here, we report a direct, one-step exosome sampling technology, ExoPRIME, for selective capture of CD63+ exosome subpopulations using an immune-affinity protocol. Microneedles (300μm × 30 mm), functionalized with anti-CD63 antibodies, were incubated under various experimental conditions in conditioned astrocyte medium and astrocyte-derived exosome suspension. The probe's capture efficiency and specificity were validated using FluoroCet assay, immunofluorescent imaging, and OMICS analyses. Significantly higher exosomes were captured by probes incubated for 16 h at 4 0C in enriched exosomal suspension (23 × 10 6 exosomes per probe) vis-à-vis 2 h at 4 0 C (12 × 10 6) and 16 h at 22 0C (3 × 10 6) in conditioned cell media. Our results demonstrate the application of ExoPRIME over a broad dynamic range of temperature and incubation parameters, offering flexibility for any desired application. ExoPRIME permits the use and re-use of minimal sample volumes (≤200 μL), can be multiplexed in arrays, and integrated into a lab-on-a-chip platform to achieve parallel, high-throughput isolation of different exosome classes in a semi-automated workstation. This platform could provide direct exosomal analysis of biological fluids since it can elegantly interface with existing room-temperature, picomolar-range nucleic acid assays to provide a clinical diagnostic tool at the point of care.