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Staphylococcal enterotoxin B (SEB) is a potent bacterial toxin that causes inflammatory stimulation and toxic shock, thus it is necessary to detect SEB in food and environmental samples. Here, we developed a sensitive immunodetection system using monoclonal antibodies (mAbs). Our study is the first to employ a baculovirus expression vector system (BEVS) to produce recombinant wild-type SEB. BEVS facilitated high-quantity and pure SEB production from suspension-cultured insect cells, and the SEB produced was characterized by mass spectrometry analysis. The SEB was stable at 4 °C for at least 2 years, maintaining its purity, and was further utilized for mouse immunization to generate mAbs. An optimal pair of mAbs non-competitive to SEB was selected for sandwich enzyme-linked immunosorbent assay-based immunodetection. The limit of detection of the immunodetection method was 0.38 ng/mL. Moreover, it displayed higher sensitivity in detecting SEB than commercially available immunodetection kits and retained detectability in various matrices and S. aureus culture supernatants. Thus, the results indicate that BEVS is useful for producing pure recombinant SEB with its natural immunogenic property in high yield, and that the developed immunodetection assay is reliable and sensitive for routine identification of SEB in various samples, including foods.

Tularemia, caused by the bacterium , poses health risks to humans and can spread through a variety of routes. It has also been classified as a Tier 1 Select agent by the CDC, highlighting its potential as a bioterrorism agent. Moreover, it is difficult to diagnose in a timely fashion, owing to the non-specific nature of tularemia infections. Rapid, sensitive, and accurate detection methods are required to reduce mortality rates. We aimed to develop antibodies directed against the outer membrane protein A of (FopA) for rapid and accurate diagnosis of tularemia. We used a baculovirus insect cell expression vector system to produce the FopA antigen and generate anti-FopA antibodies through immunization of BALB/c mice. We then employed hybridoma and phage display technologies to screen for antibodies that could recognize unique epitopes on FopA. Two monoclonal antibodies, 6B12 and 3C1, identified through phage display screening specifically bound to recombinant FopA in a dose-dependent manner. The binding affinity of the anti-FopA 6B12 and 3C1 antibodies was observed to have an equilibrium dissociation constant of 1.76 × 10-10 M and 1.32 × 10-9 M, respectively. These antibodies were used to develop a sandwich ELISA system for the diagnosis of tularemia. This assay was found to be highly specific and sensitive, with detection limits ranging from 0.062 ng/mL in PBS to 0.064 ng/mL in skim milk matrices. Our findings demonstrate the feasibility of a novel diagnostic approach for detecting based on targeting FopA, as opposed to existing tests that target the bacterial lipopolysaccharide.

Gallbladder carcinoma (GBC) exhibits poor prognosis due to local recurrence, metastasis, and resistance to targeted therapies. Using clinicopathological analyses of GBC patients along with molecular in vitro and tumor in vivo analysis of GBC cells, we showed that reduction of Dsg2 expression was highly associated with higher T stage, more perineural, and lymphatic invasion. Dsg2-depleted GBC cells exhibited significantly enhanced proliferation, migration, and invasiveness in vitro and tumor growth and metastasis in vivo through Src-mediated signaling activation. Interestingly, Dsg2 binding inhibited Src activation, whereas its loss activated cSrc-mediated EGFR plasma membrane clearance and cytoplasmic localization, which was associated with acquired EGFR-targeted therapy resistance and decreased overall survival. Inhibition of Src activity by dasatinib enhanced therapeutic response to anti-EGFR therapy. Dsg2 status can help stratify predicted patient response to anti-EGFR therapy and Src inhibition could be a promising strategy to improve the clinical efficacy of EGFR-targeted therapy.

Introduction: The binary PirA/PirB toxin expressed by Vibrio parahaemolyticus (PirABVp) is a virulent complex that causes acute hepatopancreatic necrosis disease (AHPND) in shrimps, affecting the global shrimp farming industry. AHPND is currently diagnosed by detecting pirA and pirB genes by PCR; however, several V. parahaemolyticus strains do not produce the two toxins as proteins. Thus, an immunoassay using antibodies may be the most effective tool for detecting toxin molecules. In this study, we report a sandwich ELISA-based immunoassay for the detection of PirABVp.Methods: We utilized a single-chain variable fragment (scFv) antibody library to select scFvs against the PirA or PirB subunits. Phage display panning rounds were conducted to screen and identify scFv antibodies directed against each recombinant toxin subunit. Selected scFvs were converted into IgGs to develop a sandwich immunoassay to detect recombinant and bacterial PirABVp.Results: Antibodies produced as IgG forms showed sub-nanomolar to nanomolar affinities (KD), and a pair of anti-PirA antibody as a capture and anti-PirB antibody as a detector showed a limit of detection of 201.7 ng/mL for recombinant PirABVp. The developed immunoassay detected PirABVp in the protein lysates of AHPND-causing V. parahaemolyticus (VpAHPND) and showed a significant detectability in moribund or dead shrimp infected with a VpAHPND virulent strain compared to that in non-infected shrimp.Discussion: These results indicate that the developed immunoassay is a reliable method for diagnosing AHPND by detecting PirABVp at the protein level and could be further utilized to accurately determine the virulence of extant or newly identified VpAHPND in the global shrimp culture industry.

Cancer stem-like cells (CSCs) can generate solid tumors through the properties of stem cells such as self-renewal and differentiation and they cause drug resistance, metastasis and recurrence. Therefore, establishing CSC lines is necessary to conduct various studies such as on the identification of CSC origin and specific targeted therapies. In this study, we stimulated NIH3T3 fibroblasts to exhibit the characteristics of CSCs using the whole protein lysates of B16F10 melanoma cells. As a result, we induced colony formation that displayed self-renewal and differentiation capacities through anchorage-independent culture and re-attached culture. Moreover, colonies showed drug resistance by being maintained in the G0/G1 state. Colonies expressed various CSC markers and displayed high-level drug efflux capacity. Additionally, colonies clearly demonstrated tumorigenic ability by forming a solid tumor in vivo . These results show that proteins of cancer cells could transform normal cells into CSCs by increasing expression of CSC markers. This study argues the tremendous importance of the extracellular microenvironmental effect on the generation of CSCs. It also provides a simple experimental method for deriving CSCs that could be based on the development of targeted therapy techniques.

The binary PirA/PirB toxin expressed by (PirAB ) is a virulent complex that causes acute hepatopancreatic necrosis disease (AHPND) in shrimps, affecting the global shrimp farming industry. AHPND is currently diagnosed by detecting and genes by PCR; however, several strains do not produce the two toxins as proteins. Thus, an immunoassay using antibodies may be the most effective tool for detecting toxin molecules. In this study, we report a sandwich ELISA-based immunoassay for the detection of PirAB . We utilized a single-chain variable fragment (scFv) antibody library to select scFvs against the PirA or PirB subunits. Phage display panning rounds were conducted to screen and identify scFv antibodies directed against each recombinant toxin subunit. Selected scFvs were converted into IgGs to develop a sandwich immunoassay to detect recombinant and bacterial PirAB . Antibodies produced as IgG forms showed sub-nanomolar to nanomolar affinities (K ), and a pair of anti-PirA antibody as a capture and anti-PirB antibody as a detector showed a limit of detection of 201.7 ng/mL for recombinant PirAB . The developed immunoassay detected PirAB in the protein lysates of AHPND-causing and showed a significant detectability in moribund or dead shrimp infected with a virulent strain compared to that in non-infected shrimp. These results indicate that the developed immunoassay is a reliable method for diagnosing AHPND by detecting PirAB at the protein level and could be further utilized to accurately determine the virulence of extant or newly identified in the global shrimp culture industry.

Lithium metal has shown a lot of promise for use as an anode material in rechargeable batteries owing to its high theoretical capacity. However, it does not meet the cycle life and safety requirements of rechargeable batteries owing to electrolyte decomposition and dendrite formation on the surfaces of the lithium anodes during electrochemical cycling. Here, we propose a novel electrolyte system that is relatively stable against lithium metal and mitigates dendritic growth. Systematic design methods that combined simulations, model-based experiments and in situ analyses were employed to design the system. The reduction potential of the solvent, the size of the salt anions and the viscosity of the electrolyte were found to be critical parameters determining the rate of dendritic growth. A lithium metal anode in contact with the designed electrolyte exhibited remarkable cyclability (more than 100 cycles) at a high areal capacity of 12 mAh cm −2 .

Efficient electrocatalysis at the cathode is essential for overcoming the limitations of LiO2 batteries such as poor stability and low rate capability. Herein, we systematically studied the kinetic behavior of a LiO2 battery comprising perovskite La0.8Sr0.2VO3 nanofibers formed by partial Sr‐cation doping and V cations with multiple oxidation states. Compared with undoped LaVO3 and La0.8Sr0.2VO4 nanofibers, perovskite La0.8Sr0.2VO3 nanofibers exhibited an improved capacity of 2000 mA g−1, and a 20‐times‐longer cycle life in LiO2 batteries. X‐ray photoelectron spectroscopy, electron paramagnetic resonance spectroscopy, and photoluminescence analyses revealed that the performance variations mainly originated from crystal defects, which modulate oxygen reduction/evolution kinetics. Through in situ Raman analysis, we showed that these structural defects are closely related to the oxygen reduction/evolution behavior of La0.8Sr0.2VO3 nanofibers and result in fewer parasitic reactions. This study offers insights into the potential rate capability of LiO2 batteries and related devices. Perovskite Sr‐doped LaVO3 nanofibers are prepared as a high‐rate LiO2 battery electrocatalyst for the first time. We leverage the unique properties of the perovskite structure by controlling the partial doping of Sr cations and the distribution of V cations with multiple oxidation states. La0.8Sr0.2VO3 nanofibers exhibit a high reversibility for the oxygen reduction/evolution reaction and a high‐rate capability.

To improve the maneuverability and agility of jumping robots, a variety of steerable jumping mechanisms have been actively studied. The steering ability enables a robot to reach a particular target by altering its jumping direction. To make this possible, we propose a miniature steerable jumping robot based on froghopper’s jumping principle: Moment cancellation is achieved via synchronous leg rotation, and a predictable jumping direction is achieved through an almost zero stiffness femoro-tibial joint. To satisfy these working principles, the robot is designed to have a four-bar shaped body structure and wire-driven knee joints. The four-bar body always synchronizes the leg operation by mechanically coupling the two jumping legs, which enables the robot to cancel out the moments and finally reduce the needless body spin. The knee joints are actuated using wires, and the wires are kept loose to maintain joint stiffness almost zero during take-off. Accordingly, the jumping direction is successfully predicted to determine the initial posture of the tibia. As a result, the proposed robot can change the jumping direction from −20 degrees to 20 degrees while reducing needless body spin.

With the development of nanotechnology, myriad types of novel materials have been discovered at the nanoscale, among which the most interesting material is graphene. However, the toxicity data available on graphene are extremely limited. In this study, we explored toxic response of commercially available graphene nanoplatelets (GNPs) in vivo and in vitro. The GNPs used in this study had a high surface area and feature considerably few defects. In mice, GNPs (2.5 and 5 mg/kg) remained in the lung until 28 days after a single instillation, and the secretion of inflammatory cytokines reached the maximal level at Day 14 and then decreased over time. In vitro study using BEAS-2B cells, a human bronchial epithelial cell line, GNPs located within autophagosome-like vacuoles 24 h after exposure. The GNPs (2.5, 5, 10, and 20 μg/mL) also dose-dependently reduced cell viability, which was accompanied by an increase in the portion of cells in the subG1 and S phases. Moreover, the GNPs down-regulated the generation of reactive oxygen species, suppressed ATP production, caused mitochondria damage, and elevated the levels of autophagy-related proteins. Based on these results, we suggest that GNPs provoked a subchronic inflammatory response in mice and that GNPs induced autophagy accompanying apoptosis via mitochondria damage in vitro.