Cell Preparations Research Articles

ATP-Gated P2X7-Ion Channel on Kidney-Resident Natural Killer T Cells and Memory T Cells in Intrarenal Inflammation.

The P2X7 ion channel, a key sensor of sterile inflammation, has been implicated as a therapeutic target in glomerulonephritis, and P2X7-antagonistic nanobodies can attenuate experimental glomerulonephritis. However, little is known about the expression of P2X7 on renal immune cells. We used conventional immunofluorescence of kidney sections as well as intraperitoneal injection of nanobodies in mice followed by flow cytometry analysis of parenchymal T cells and RNA-sequencing to elucidate the expression and function of P2X7 on parenchymal and vascular immune cells in the mouse kidney. Our study showed that parenchymal T cells, including a large subset of natural killer T cells and CD69+ tissue-resident memory T cells, display much higher cell surface levels of P2X7 than vascular T cells. After a single intraperitoneal injection of P2X7-blocking nanobodies, P2X7 on parenchymal T cells was fully occupied by the injected nanobodies within 30 min. This resulted in an effective protection of these cells from NAD-induced cell death during cell preparation. Conversely, systemic injection of NAD that mimics sterile inflammation results in the selective depletion of P2X7hiCD69hi T cells from the kidney parenchyma. Our study uncovered a novel purinergic regulatory mechanism affecting kidney-resident T cell populations.

Multi functional ionic Ru(bpy)3(PF6)2 assisted preparation of perovskite solar cell with over 19% and superior stability

Abstract The global community is striving to advance the development of emerging perovskite solar cells (PSCs). It has been demonstrated that the utilization of passivators is an effective approach to enhance the photoelectric conversion efficiency and long-term stability of PSCs, thus attracting increasing attention to organic passivators. All-inorganic CsPbI3 PSCs play a crucial role in the next-generation photovoltaic technology due to their unique optical bandgap and excellent light absorption properties. However, due to its poor stability, CsPbI3−x Br x halide hybrid perovskite has been proposed. The spin-coating film formation method leads to the presence of a large number of vacancy defects in the film, which poses a significant challenge to the commercialization of CsPbI3−x Br x perovskite films. Therefore, in this study, a novel passivator, Ru(bpy)3(PF6)2 (TRH), was introduced. By introducing the passivator, the photoelectric conversion efficiency of PSCs in air was improved, there by optimizing the device performance. After the addition of the passivator, [Ru(bpy)3]2+ filled the defects on the surface of the film, while the free PF6 - ions passivated the negatively charged halide vacancies. The fluorine atoms effectively prevented the entry of water molecules in the air. As a result, the optimized CsPbI3−x Br x film exhibited a more compact morphology, a lower defect state density, and better carrier transport performance. Eventually, the champion device achieved a photoelectric conversion efficiency of 19.47%, and even after being placed at an ambient temperature of 25 °C–30°C and a humidity of 30%–40% for 150 d, its efficiency remained above 85% of the initial efficiency of the device.

Intra-Articular Injection of Stem Cells for the Regeneration of Knee Joint Cartilage: a Therapeutic Option for Knee Osteoarthritis - a Narrative Review.

Current approaches to regulating osteoarthritis primarily focus on symptom management; however, these methods often have significant side effects and may not be suitable for long-term care. As an alternative to conventional treatments, injecting stem cells into knee joint cartilage is a promising option for repairing damaged cartilage. In this review, we outline the general procedure for stem cell treatment of knee joint cartilage regeneration, emphasizing the potential of intra-articular stem cell injections as a therapeutic option for osteoarthritis. We examined and summarized patient evaluation and preparation for knee joint stem cell therapy, stem cell harvesting, stem cell preparation, injection procedures for stem cell therapy, post-injection care and monitoring, potential outcomes of stem cell therapy, and considerations and risks associated with stem cell therapy. Overall, stem cell injections for knee joint cartilage damage represent a promising frontier in orthopedic care. They offer potential benefits such as pain and inflammation reduction, promotion of cartilage repair and regeneration, and the possibility of avoiding more invasive treatments such as knee surgery. Ongoing collaboration among researchers, clinicians, and regulatory organizations is crucial for advancing this field and translating scientific discoveries into effective clinical applications.

Evaluation of the immune status of dogs vaccinated against rabies by an enzyme-linked immunosorbent assay using crude preparations of insect cells infected with a recombinant baculovirus encoding the rabies virus glycoprotein gene.

Evaluation of the effectiveness of vaccination of animals against rabies is not routinely implemented. In cases where it is carried out, the rapid fluorescent focus inhibition test (RFFIT) or the fluorescent antibody virus neutralization (FAVN) test are the recommended tests. However, both of these tests require handling of live rabies virus (RABV), and are cumbersome to perform. In view of this, the enzyme-linked immunosorbent assay (ELISA) has been proposed as a surrogate test; however, availability of appropriate antigen is a major impediment for the development of ELISAs to detect anti-rabies antibodies. The most widely used antigen is the RABV glycoprotein (G) purified from cell culture-propagated virus, which requires a biosafety level 3 containment. The alternative is to use recombinantly expressed G, which needs to be to be properly glycosylated and folded to serve as the best antigen. The most suitable system for its production is the baculovirus expression system (BVES). However, purification of RABV G is challenging. We therefore tested partially purified preparations in the form of extracts of insect cells infected with baculovirus expressing RABV G, against sera from vaccinated dogs in an indirect ELISA. The results showed good concordance against RFFIT, with sensitivity and specificity of 90.48% and 80.00%, respectively. The system may be used for quick screening to determine the presence and an approximate level of antibodies, and can be modified to enable monitoring of mass dog vaccination programs, as well as to facilitate certification of dogs intended for international travel and transportation.

Cell Division Imaging of Tobacco BY-2 Cells in a 3D Volume by Two-Photon Spinning-Disk Confocal Microscopy.

Plant cells have a unique organization of microtubules during mitosis and cytokinesis. However, it is difficult to observe cell division in plant cells because of the distortion of images caused by thick samples. We addressed this issue by two-photon excitation coupled with spinning-disk confocal microscopy. Herein, we describe the method for observing microtubules and nuclei during cell division in tobacco BY-2 cells by two-photon spinning-disk confocal microscopy. The protocol includes the transformation of BY-2 cells with a plasmid for the expression of tubulin and histone markers, preparation of cells for microscopy, and operation of the two-photon spinning-disk microscope.

A dendritic cell vaccine for both vaccination and neoantigen-reactive T cell preparation for cancer immunotherapy in mice

Adoptive cell transfer (ACT) using neoantigen-specific T cells is an effective immunotherapeutic strategy. However, the difficult isolation of neoantigen-specific T cells limits the clinical application of ACT. Here, we propose a method to prepare neoantigen-reactive T cells (NRT) for ACT following immunization with a tumor lysate-loaded dendritic cell (DC) vaccine. We show that the DC vaccine not only induces a neoantigen-reactive immune response in lung cancer-bearing mice in vivo, but also facilitate NRT cell preparation in vitro. Adoptive transfer of the NRTs as combinatorial therapy into DC vaccine-immunized, LL/2 tumor-bearing mice allows infiltration of the infused NRTs, as well as the enrichment of neoantigen reactive, non-ACT/NRT T cells into the tumor microenvironment with the function of these neoantigen-reactive T-cell receptors validated in vitro. In summary, we propose a method for preparing NRTs that increases ACT efficacy and paves the way to the design of personalized immunotherapies.

Further mobility enhancements for 5G-advanced and beyond

The study on mobility is an inherent part of any generation of cellular technology. It is aimed at ensuring robust, reliable, and interruption-free handover of the mobile user connection. Such continuous improvement is necessary as the service requirements become more stringent which urge the mobility to meet the highest performance expectations. In this paper, it is described how such requirements are to be achieved by the 3GPP Release 18 components. It begins with the introduction to the world of mobility, covering the solutions that have been designed since the first generation of New Radio (NR). Then improvements pursued in consecutive 3GPP releases are outlined. Selected areas for Release 18 mobility enhancements have been evaluated using 5G-compliant system-level simulations and results thereof are presented below. These include wrong Primary Secondary Cell (PSCell) preparation in Conditional Handover (CHO) and the impacts of secondary cell setup delay in Dual Connectivity (DC). It is shown that preparing multiple PSCells in CHO ensures the user accesses the right cell in up to 96% of cases.

Exploring Sb2S3 as a hole transport layer for GeSe based solar cell: a numerical simulation

Abstract Germanium selenide (GeSe) and antimony sulfide (Sb2S3) both are technological intriguing semiconductor material for green and economical photovoltaic devices. In this study, GeSe and Sb2S3 have been utilized as the absorber layer and hole transport layer, respectively, to constructed a heterojunction thin film solar cell consisting of FTO/TiO2/GeSe/Sb2S3/Metal. The GeSe and Sb2S3 are binary compounds and can adopt the same film deposition method, for instance, thermal evaporation, which is expected to improve process compatibility and to reduce production costs. The TiO2 (electron transport layer) and Sb2S3 can form small spike-like conduction band offset and valence band offset with the GeSe, respectively, which possesses potential to suppress carrier recombination at the heterointerfaces. Subsequently, the effects of main functional layer material parameters, heterointerface characteristics and back contact metal work function on the performance parameters of the proposed solar cell were simulated and analyzed using wxAMPS software. After numerical simulation and optimization, the proposed solar cell can reach an open circuit voltage of 0.872 V, a short circuit current of 40.72 mA·cm−2, a filling factor of 84.16%, and a conversion efficiency of 28.35%. According to the simulation results, it is anticipated that the Sb2S3 can serve as a hole transport layer for GeSe based solar cell and enable device to achieve high efficiency. Simulation analysis also provides some meaningful references for the design and preparation of heterojunction thin film solar cells.

Review and Practical Guide for Getting Started With Single-Cell Proteomics.

Single-cell proteomics (SCP) has advanced significantly in recent years, with new tools specifically designed for the preparation and analysis of single cells now commercially available to researchers. The field is sufficiently mature to be broadly accessible to any lab capable of isolating single cells and performing bulk-scale proteomic analyses. In this review, we highlight recent work in the SCP field that has significantly lowered the barrier to entry, thus providing a practical guide for those who are newly entering the SCP field. We outline the fundamental principles and report multiple paths to accomplish the key steps of a successful SCP experiment including sample preparation, separation, and mass spectrometry data acquisition and analysis. We recommend that researchers start with a label-free SCP workflow, as achieving high-quality and quantitatively accurate results is more straightforward than label-based multiplexed strategies. By leveraging these accessible means, researchers can confidently perform SCP experiments and make meaningful discoveries at the single-cell level.

In Situ Tumor-Infiltrating Lymphocyte Therapy by Local Delivery of an mRNA Encoding Membrane-Anchored Anti-CD3 Single-Chain Variable Fragment.

Tumor-infiltrating lymphocyte (TIL) therapy has shown promising responses in clinical trials for highly aggressive cancers such as advanced melanoma and metastatic colorectal cancer. However, TIL therapy is still limited in clinical practice due to the complex ex vivo cell preparation process. Here, we report an "in situ TIL therapy" for the treatment of solid tumors. We utilized lipid nanoparticles for the delivery of an mRNA encoding membrane-anchored anti-CD3 single-chain variable fragment (scFv) (MA-aCD3), efficiently engineering both tumor-associated macrophages (TAMs) and tumor cells following intratumoral delivery. Expression of MA-aCD3 resulted in enhanced TIL activation, proliferation, and tumor cell engagement directly within the tumor microenvironment. In B16F10 and MC38 tumor models, concurrent expression of MA-aCD3 on TAMs and tumor cells mediated by mRNA delivery resulted in significant antitumor effects via in situ polyclonal CD8+ TIL expansion and directed cytotoxic effector functions. In addition, combinatorial treatment of MA-aCD3-encoding mRNA and antiprogrammed cell death 1 (anti-PD-1) antibodies exhibited synergistic antitumor effects on anti-PD-1 refractory B16F10 tumors. Together, our findings suggest that in situ TIL therapy is a practical and effective mRNA-based therapeutic modality for the treatment of solid tumors.

Preparation of efficient all-inorganic perovskite solar cells from large grain via 1-heptanamine stabilised CsPbI2Br nanoparticles with NH4SCN additive

CsPbI2Br is a potential material for perovskite solar cells (PSCs) due to its better thermal stability and high efficiency. Centrifugal deposition for film formation greatly simplifies the steps of the conventional anti-solvent spin coating method, but there are still problems hindering progress, such as small grain size and poor photoelectric performance. In this work, NH4SCN was introduced as an additive during the preparation of heptylamine (C7H17N) stabilised CsPbI2Br nanoparticles (NCs) and the films were prepared by centrifugal deposition. The results show that the low melting temperature compounds which formed by thiourea from NH4SCN decomposition during annealing and unreacted PbX2 would link the small grains together by melting and flowing; after PbX2 precipitation and reaction with CsX, the grain size was significantly increased by about 95 % compared to that of the untreated film. Due to large grains, less defects and high crystallinity, the final power conversion efficiency and fill factor reached 12.25 % and 0.70, respectively.

Studying the effect of ambient temperature variations on perovskite precursor film formation using different antisolvents

This study investigates the sensitivity of two antisolvents to ambient temperature fluctuations during the fabrication of triple-cation perovskite thin films. Chlorobenzene (CB) and ethyl acetate (EA) were employed as antisolvents in the preparation of perovskite solar cells (PSCs) under identical ambient temperature variation conditions. The experimental results reveal that the quality of the perovskite films does not exhibit a consistent trend in response to ambient temperature changes depending on the antisolvent used. When CB was used as the antisolvent, the highest-performing device, produced at an ambient temperature of 18 °C, achieved a power conversion efficiency (PCE) of 19.9 %. In contrast, when EA was employed as the antisolvent, the highest-performing device was fabricated at 25 °C, reaching a PCE of 20.5 %. Furthermore, when the ambient temperature exceeded 30 °C, all films prepared with either antisolvent exhibited significant cracking, indicating that elevated temperatures adversely affect perovskite film formation. To further investigate the mechanism through which ambient temperature affects the film-forming process, antisolvents at varying temperatures were used during perovskite film preparation to simulate transient temperature increases during formation. This study reveals that the impact of ambient temperature variations on perovskite film formation does not exhibit a uniform trend but is significantly influenced by the choice of antisolvent. Moreover, this work offers valuable insights for optimizing the fabrication of high-quality perovskite thin films.

Generation of Antibody Libraries for Phage Display: Human Fab Format.

Phage display is a powerful method for the de novo generation and affinity maturation of human monoclonal antibodies from naive, immune, and synthetic antibody repertoires. The pComb3 phagemid family of phage display vectors facilitates the selection of human monoclonal antibody libraries in the monovalent Fab format, which consists of human variable domains VH and VL (Vκ or Vλ), fused to the human constant domains CH1 of IgG1 and CL (Cκ or Cλ), respectively. Here, we describe the use of a pComb3 derivative, phagemid pC3C, for the generation of human Fab libraries with randomly combined human variable domains (VH, Vκ, and Vλ) of high sequence diversity, starting from the preparation of mononuclear cells from blood and bone marrow. Depending on the complexity of the parental antibody repertoire, the protocol can be scaled for yielding a library size of 108-1011 independent human Fab clones. As such, it can be used, for instance, for the generation of a large naive human Fab library from healthy individuals or for the generation of a specialized immune human Fab library from individuals with an endogenous antibody response of interest.

11.88% Efficient Flexible Ag-Free CZTSSe Solar Cell: Spontaneously Tailoring the Alkali Metal Level.

Alkali metal is the requirement for highly efficient Cu2ZnSn(S, Se)4 (CZTSSe) solar cells, thus it is crucial to additionally incorporate alkali metal into the absorber layer for flexible solar cells. However, the efficiency of flexible CZTSSe devices reported to date, based on the conventional alkali incorporation strategies, still lags behind those made on rigid substrates. One of the main issues is the inability to control the alkali content and distribution in the absorber layer. Here, a facile alkaline incorporation approach is proposed, effectively regulating the content and distribution of alkali metals in the film. Such a method can spontaneously tailor the alkali metal content to a proper level, thus leading to the suppression of non-radiative recombination and a better carrier transport through the enhanced film quality and the optimized band binding structure. Finally, a champion flexible CZTSSe solar cell with an efficiency of 11.88% is achieved, the highest reported efficiency for a CZTSSe solar cell without noble Ag doping. This study affords an innovative spontaneous alkali-doping design for the preparation of high-performance flexible CZTSSe solar cells and provides a deeper insight into the extent of alkali metal doping.

Thermo-responsible PNIPAM-grafted polystyrene microspheres for mesenchymal stem cells culture and detachment.

The preparation of cells is a critical step in cell therapy. To ensure the effectiveness of cells used for clinical treatments, it is essential to harvest adherent cells from the culture media in a way that preserves their high viability and full functionality. In this study, we developed temperature-responsive poly(N-isopropylacrylamide) (PNIPAM)-grafted polystyrene (PS) microspheres using reversible addition-fragmentation chain transfer polymerization. These microspheres allow for the non-destructive harvesting of cultured cells through temperature changes. The composition and physicochemical properties of the PNIPAM-grafted PS microspheres were confirmed using infrared spectroscopy, elemental analysis, dynamic light scattering, and thermogravimetric analysis.In vitroexperiments demonstrated that these microspheres exhibit excellent biocompatibility, supporting the adhesion and proliferation of various cells. Moreover, the microspheres showed good temperature responsiveness in thermosensitive detachment experiments with GFP-HepG2cells and umbilical cord mesenchymal stem cells (UC-MSCs). Additionally, through orthogonal experiments, we identified a cell detachment aid mixture that significantly improved the dispersibility of cells detached from the microspheres, enhancing the efficiency of thermosensitive cell detachment by approximately 40%. The harvested UC-MSCs retained their capacity for re-proliferation and trilineage differentiation. Consequently, the temperature-responsive microspheres developed in this study, combined with the cell detachment aid mixtures, hold great potential for large-scale culture and harvesting of therapeutic cells in clinical applications.

Reviving Sodium Tunnel Oxide Cathodes Based on Structural Modulation and Sodium Compensation Strategy Toward Practical Sodium-Ion Cylindrical Battery.

As a typical tunnel oxide, Na0.44MnO2 features excellent electrochemical performance and outstanding structural stability, making it a promising cathode for sodium-ion batteries (SIBs). However, it suffers from undesirable challenges such as surface residual alkali, multiple voltage plateaus, and low initial charge specific capacity. Herein, an internal and external synergistic modulation strategy is adopted by replacing part of the Mn with Ti to optimize the bulk phase and construct a Ti-containing epitaxial stabilization layer, resulting in reduced surface residual alkali, excellent Na+ transport kinetics and improved water/air stability. Specifically, the Na0.44Mn0.85Ti0.15O2 using water-soluble carboxymethyl cellulose as a binder can realize a capacity retention rate of 94.30% after 1,000 cycles at 2C, and excellent stability is further verified in kilogram large-up applications. In addition, taking advantage of the rich Na content in Prussian blue analog (PBA), PBA-Na0.44Mn1-xTixO2 composites are designed to compensate for the insufficient Na in the tunnel oxide and are matched with hard carbon to achieve the preparation of coin full cell and 18650 cylindrical battery with satisfactory electrochemical performance. This work enables the application of tunnel oxides cathode for SIBs in 18650 cylindrical batteries for the first time and promotes the commercialization of SIBs.

Unveiling the role of Cs ion in perovskite phase formation during solid–vapor reactions

The preparation of perovskite solar cells (PSCs) using a solid–vapor reaction has shown great potential due to its scalability and lack of organic solvent. The phase formation process in the solid–vapor reaction is crucial for the crystallization of perovskite films. Previous studies mainly introduced Cs into the solid–vapor reaction, but insufficient attention was paid to the role of Cs in purifying the phase formation path during the crystallization process. In this study, we investigated the effect of Cs on the phase formation process in the solid–vapor reaction and found that Cs can decrease the crystallinity of lead halide films, induce a purified phase formation process to prevent the formation of δ-phase and other impurity phases, regulate the crystallization rate of perovskite, and produce high-quality, low-defect perovskite films. The result was achieving a champion power conversion efficiency (PCE) of 21.54 % for a small area (0.148 cm2) and 18.65 % for larger mini-modules (5 × 5 cm2). Furthermore, the devices maintained over 80 % of their initial efficiencies after 450 h of continuous operation under AM 1.5 G irradiation. This work underscores the importance of purified phase formation paths in solid–vapor reactions for outstanding crystal quality in high-performance perovskite solar cells.

Evidencing Phase Transformations of Li6PS5Cl Argyrodite under Ambient Air and Dry Room Exposure.

All-solid-state electrolytes have been extensively studied for the last years in order to achieve high conductivities and improved safety among lithium-ion technologies. Sulfide electrolytes, such as argyrodites (Li6PS5X, X = Cl, Br, and I), succeed to show high performances despite their poor chemical stability. As a matter of fact, argyrodite reactivity to water is known as a common drawback for easy implementation and requires the use of dry room for cell preparation. The understanding of argyrodite degradation under ambient air exposure is a key for the development of stable electrolytes, coatings, and processes and has been incompletely explored until now. This study brings unreported elements of comprehension around the degradation mechanisms of Li6PS5Cl solid electrolyte using transmission electron microscopy (TEM) and complementary spectroscopic techniques.

The effects of time and anticoagulant type on hematologic values in cockatiels (Nymphicus hollandicus)

BackgroundAvian hematologic characteristics complicate evaluation; therefore, complete blood cell counts require manual analysis for best accuracy. Furthermore, the quality of samples may be affected by prolonged shipment. MethodsBlood was collected from the right jugular vein of twenty-four, 7-year-old (14 male, 10 female), wild-type cockatiels (Nymphicus hollandicus). Smears were prepared using fresh, untreated blood. The remainder was divided into dipotassium ethylenediaminetetraacetic acid (EDTA) and lithium heparin microtainers. A hematocrit tube and blood smear were prepared at baseline, 24, 48, and 72 hours after venipuncture. Packed cell volume (PCV), total solids (TS), estimated white blood cell (WBC) count, WBC differential (heterophils, lymphocytes, monocytes, eosinophils, basophils), hemolysis index (HI), and preparation quality were assessed and compared to baseline. After log transformation, the effects of anticoagulants across time points were compared. ResultsHeparinized samples had significant differences in WBC, heterophil, and lymphocyte counts at 48 hours, and PCV and TS at 72 hours compared to baseline. There was a positive proportional bias in PCV and TS from EDTA anticoagulant samples at 24, 48, and 72 hours, and from heparinized samples at 48 and 72 hours compared to baseline. The EDTA samples had increased PCV and TS compared to heparin at all time points, and higher WBC and heterophil counts at 48 hours compared to baseline. A type II methodological error cannot be excluded. ConclusionThese results suggest that the anticoagulant used and time spent in anticoagulant alters hematologic values in cockatiels, which may have significant clinical implications. The importance of sample processing within 24 hours and anticoagulant-specific reference intervals for hematologic variables should be considered.

RNA Sequencing of Sperm from Healthy Cattle and Horses Reveals the Presence of a Large Bacterial Population.

RNA molecules within ejaculated sperm can be characterized through whole-transcriptome sequencing, enabling the identification of pivotal transcripts that may influence reproductive success. However, the profiling of sperm transcriptomes through next-generation sequencing has several limitations impairing the identification of functional transcripts. In this study, we explored the nature of the RNA sequences present in the sperm transcriptome of two livestock species, cattle and horses, using RNA sequencing (RNA-seq) technology. Through processing of transcriptomic data derived from bovine and equine sperm cell preparations, low mapping rates to the reference genomes were observed, mainly attributed to the presence of ribosomal RNA and bacteria in sperm samples, which led to a reduced sequencing depth of RNAs of interest. To explore the presence of bacteria, we aligned the unmapped reads to a complete database of bacterial genomes and identified bacteria-associated transcripts which were characterized. This analysis examines the limitations associated with sperm transcriptome profiling by reporting the nature of the RNA sequences among which bacterial RNA was found. These findings can aid researchers in understanding spermatozoal RNA-seq data and pave the way for the identification of molecular markers of sperm performance.