Search Results (742)

Celiac patients require strict avoidance of gliadin, the primary immunotoxic component of gluten, making sensitive detection essential for food safety. A label-free and reagentless capacitive aptasensor for gliadin detection was developed using an aminated polyethylene terephthalate (PET) thin film as both an insulating layer and functionalization platform. The PET surface was modified via ethylenediamine-mediated aminolysis, enabling covalent immobilization of 5′-NH₂-modified gliadin aptamers through glutaraldehyde crosslinking. Under optimized conditions (23 µm initial PET thickness and 10 µM aptamer), the sensor showed a linear response from 10 to 500 µg/mL gliadin (R² = 0.9792), with a detection limit of 6.0 µg/mL, equivalent to 12 ppm gluten, which is well below the regulatory threshold of 20 ppm for gluten-free labeling. The aptasensor showed excellent correlation with commercial ELISA for 20 gluten-containing soy sauce samples (R² = 0.926) and spike recoveries of 91.7–105.7% in two gluten-free products. Efficient regeneration was achieved with 25 mM arginine (pH 9.0), retaining >80% activity after six cycles. This simple, low-cost, and reusable platform relies solely on a single PET thin film as consumable in a custom-built system with lab-friendly aminolysis conditions. It substantially lowers barriers to functionalized insulating layer fabrication, the primary challenge in capacitive aptasensor development, providing a promising method for on-site gliadin monitoring in gluten-free food safety applications. Full article

Objective: Effectively eliminating xenoimmunogenicity and achieving recellularization in cardiac xenografts remains a critical challenge in developing an ideal implantable xenograft. We have previously demonstrated that the removal of major antigens, including Galα1-3Gal (α-Gal) epitope and non-human sialic acid N-glycolylneuraminic acid (Neu5Gc), using α-galactosidase and peptide N-glycosidase F (PNGase-F), enables a synergistic effect with decellularization, significantly reducing the expression of carbohydrate-binding lectins without altering the biomechanical properties of the graft. The aim of this study was to establish an effective method for in vitro recellularization by seeding human mesenchymal stem cells (MSCs) on decellularized cardiac xenografts that had undergone optimal xenoantigen removal using α-galactosidase and PNGase-F. Additionally, this study aimed to evaluate the potential for in vivo recellularization. Methods: Decellularized porcine pericardium scaffolds treated with both enzymes were further modified by forming a fibrin mesh on their surface and within their structure, followed by the attachment of heparin and human vascular endothelial growth factor to the mesh. Subsequently, the scaffolds were seeded with human adipose tissue-derived stem cells for 8 weeks. In vitro recellularization, differentiation, and extracellular matrix remodeling of decellularized and enzyme-treated xenografts were assessed using vimentin, calponin, fibronectin, CD31, VWF, and phalloidin staining. To evaluate the potential for in vivo recellularization, decellularized glutaraldehyde-crosslinked xenografts with anticalcification treatments were seeded with rat bone marrow MSCs and implanted into rats subcutaneously to evaluate cell infiltration and calcification via histology, von Kossa staining, and micro-computed tomography. Results: In decellularized xenografts treated with both enzymes, stronger signals were detected and mesenchymal cell infiltration into the tissue was significantly faster, leading to accelerated recellularization. This recellularization process was more pronounced as time went on, with greater cell infiltration and evidence of cell differentiation. An in vivo study showed that decellularization and anticalcification treatments revealed stronger vimentin staining in histological analysis. The recellularization for our biocompatible scaffolds exhibited a lower degree of calcification compared to the non-recellularized tissue. Conclusions: We successfully developed major xenoantigen-free scaffolds by demonstrating the safety and synergistic effect of α-galactosidase and PNGase-F treatments and proved, for the first time, the effectiveness of recellularization using a human MSC monoculture on xenoantigen-free scaffolds. Furthermore, there was potential for in vivo recellularization of our biocompatible scaffolds seeded with MSCs. Full article

Accurate visualization of the plasma membrane is essential for assessing morphological changes induced by experimental perturbations. Scanning electron microscopy (SEM) enables high-resolution imaging of cell surface architecture, but the fixation, dehydration, and drying steps critically affect membrane preservation. Here, we present an improved SEM preparation workflow for two cultured cell models with different growth phenotypes, HepG2 and IM-9, and evaluate the effects of fixation time and drying strategy on plasma membrane preservation. Among the tested conditions, a shorter glutaraldehyde fixation time (15 min) combined with stepwise ethanol-to-HMDS substitution provided the best overall preservation of membrane continuity, cell shape, and surface regularity. Morphometric analysis supported the qualitative SEM observations, and HMDS-processed samples also showed better structural stability during storage under the tested conditions. This workflow provides a simple, reproducible, and cost-effective strategy for SEM-based analysis of cell surface morphology in cultured cell models. Full article

This study presents a headset-type biofluorometric gas sensor incorporating a CMOS camera for continuous, non-invasive monitoring of transcutaneous ethanol from the ear canal. The sensor employs alcohol dehydrogenase (ADH) to catalyze the NAD⁺-to-NADH conversion during ethanol oxidation, enabling quantitative measurement through NADH fluorescence detection (λ_ex = 340 nm, λ_em = 490 nm). The integrated system comprises a wireless CMOS camera, an ADH-immobilized cotton mesh enzyme membrane, UV-LED excitation source, optical bandpass filters, and a dual convex lens assembly housed in a 3D-printed headset powered by a lithium battery. Key improvements include a 3.5-fold enhancement in fluorescence collection efficiency achieved through optimized dual convex lens configuration. Systematic screening of seven cotton mesh materials identified Iwatsuki cotton mesh as the optimal enzyme immobilization substrate, exhibiting minimal autofluorescence and 14.2-fold higher water retention capacity compared to H-PTFE membranes. The glutaraldehyde-crosslinked ADH-immobilized cotton mesh maintained enzymatic activity for over 45 min with a 10-fold improvement in signal-to-noise ratio. The system demonstrated a dynamic detection range spanning 10 ppb to 10 ppm for gaseous ethanol and exhibited high selectivity against interfering volatile organic compounds in skin gas, including methanol, acetaldehyde, formaldehyde, and acetone. Human experiments validated the system’s practical performance. Following alcohol consumption, subjects wore the device for 50 min while real-time fluorescence monitoring captured dynamic ethanol concentration changes in the ear canal. The dose-dependent fluorescence response—approximately 2-fold higher at 0.4 g/kg versus 0.04 g/kg alcohol intake—correlated well with calibration data. This headset-type biofluorometric sensor enables unrestrained continuous monitoring of ear canal ethanol, providing a novel wearable platform for alcohol metabolism assessment with potential applications in health monitoring and clinical research. Full article

This study optimized corncob-derived magnetic biochar (MBC) with tailored iron content for laccase immobilization to boost nonylphenol (NP) degradation in water. Low-iron MBC (LMBC) and high-iron MBC (HMBC) from corncob were prepared via impregnation–pyrolysis at 700 °C, followed by modification with polyethyleneimine (PEI) and covalent immobilization of laccase using glutaraldehyde. Compared to HMBC, LMBC exhibited more mesopores, superior laccase activity recovery (60.5%) and Fe²⁺ release that was only 1/78 of HMBC’s. Furthermore, PEI modification and subsequent laccase immobilization on LMBC significantly reduced iron leaching (p < 0.05). X-ray diffraction and vibrating sample magnetometry analyses indicated that LMBC consists of crystalline Fe₃O₄, α-Fe, graphite, and carbon, with a saturation magnetization of 45.8 emu g⁻¹. Scanning electron microscopy, Fourier transform infrared spectroscopy, and X-ray photoelectron spectroscopy confirmed the microporous and mesoporous structure of LMBC, the successful PEI modification, and the effective immobilization of laccase on LMBC (L-LMBC). L-LMBC exhibited enhanced stability under acidic conditions (pH 3–7) and elevated temperatures (20–70 °C). After six repeated cycles, the NP removal efficiency by L-LMBC remained at 84.8%. Moreover, L-LMBC demonstrated effective NP removal in real environmental water samples. This study provides fundamental insights and demonstrates the potential application of MBC-immobilized laccase for degrading persistent pollutants in aqueous systems. Full article

This study aimed to develop a novel fluorescent sensor based on Eu³⁺ complex-doped protein crystal (EC-PC) for the efficient detection of metal ions in blood. By meticulously controlling the crystallization and annealing conditions in the co-crystallization strategy, the crystal growth processes were optimized to obtain doped Eu³⁺ complex-co-protein crystalline (EC-PC) structures. Thus, through co-crystallization of hen egg white lysozyme (HEWL) as a model protein and Eu³⁺ complex as fluorescent center, we successfully prepared Eu³⁺ complex-doped-HEWL co-crystals (EC-HC) with excellent fluorescent properties. Further treatment with 4% glutaraldehyde cross-linking enhanced the structural stability of the co-crystals. Moreover, the characteristic of sensitive, selective quenching of EC-PC fluorescence by biologically relevant cations, such as Cu²⁺, Zn²⁺, Mg²⁺, Ca²⁺ and Fe³⁺ ions, set up a smart sensing system in blood. For example, the fluorescence intensity of the crystals at 610 nm, as measured by a UV–visible spectrophotometer, decreases dose-dependently with the concentration of copper ions, thereby validating the sensor’s high sensitivity to copper ion detection. Significantly, we also found that this hybrid protein-based sensor did not induce hemolysis, at various volume concentrations, confirming good anticoagulation in blood. This research not only provides a new perspective on the application of Eu³⁺ complex-doped protein crystals in the field of biosensing but also offers a new strategy for the detection of biologically relevant cations in blood. Future work will focus on further optimizing the sensor’s performance and exploring its potential applications in clinical sample analysis. Full article

This study presents a highly efficient and sustainable biocatalytic platform for bisphenol A (BPA) bioremediation through the covalent immobilization of laccase onto hierarchically functionalized cotton fibers. The immobilization strategy involved selective periodate oxidation of cellulose, grafting a hexamethylenediamine (HMDA) spacer arm, and glutaraldehyde activation, ensuring stable covalent attachment. Characterization via FTIR, SEM, and BET confirmed successful surface modification and high enzyme loading, achieving an immobilization yield of 90.5%. The immobilized laccase (CT-DA-HMD-Lac) exhibited significantly enhanced performance compared to the free enzyme, with a two-fold increase in maximum reaction velocity (V_max) and a 75% improvement in catalytic efficiency of action (V_max/K_m). Furthermore, the biocatalyst demonstrated superior robustness, maintaining high activity across broader pH and temperature ranges, and retaining 75% of its initial activity after 15 consecutive reusability cycles. Storage stability was also markedly improved, with 83% activity retention after 60 days. Practical application in BPA degradation showed 85% removal efficiency within 300 min, a 2.4-fold increase in the degradation rate constant over the free enzyme. These results highlight functionalized cotton as a promising, cost-effective, and scalable support for advanced enzymatic wastewater treatment and the remediation of persistent endocrine-disrupting chemicals. Full article

This study systematically compared the induction and resuscitation characteristics of the viable but non-culturable (VBNC) state in Klebsiella pneumoniae FY170-1 following sublethal exposure to sodium hypochlorite (NaClO) or glutaraldehyde (GA). Treatment with 30 mg/L NaClO or 60 mg/L GA for 60 min reduced culturability to below the detection limit (<1 CFU/mL). However, CTC staining showed that 50.80% and 63.44% of cells, respectively, retained respiratory activity, while SYTO 9/PI staining indicated that membrane integrity was largely preserved, consistent with induction of the VBNC state. Scanning electron microscopy revealed distinct morphological alterations in the two groups. NaClO-induced VBNC cells showed surface depressions and wrinkling, consistent with oxidative damage, whereas GA-induced cells exhibited filamentous and net-like surface structures, consistent with aldehyde-mediated cross-linking. Among the tested additives, sodium succinate showed the strongest resuscitation-promoting effect under the experimental conditions, with OD600 increasing after approximately 2 h of incubation. Post-resuscitation analysis further revealed marked differences between the two VBNC states. In resuscitated NaClO-induced VBNC cells, ATP partially recovered, but reactive oxygen species remained elevated and catalase activity showed little recovery. In contrast, resuscitated GA-induced VBNC cells exhibited lower ATP recovery but more rapid normalization of ROS and better recovery of oxidative stress-related parameters. Total protein analysis and SDS-PAGE further supported distinct patterns of protein-level alteration between the two treatments. Overall, these findings suggest that NaClO and GA induce phenotypically distinct VBNC states in K. pneumoniae, with different recovery behaviors and stress response profiles. Sodium succinate was identified as the most effective recovery-promoting additive under the tested conditions. These results highlight the risk of underestimating bacterial survival when culturability is used as the sole indicator of disinfection efficacy and support the need for more comprehensive viability assessment. Full article

Fractures are becoming a bigger and bigger global health problem, with an estimated 178 million new cases each year and 455 million people living with disabilities caused by fractures. Donor site morbidity, the risk of immune rejection, and limited functional integration all make current grafting techniques less effective. Biomaterials that come from nature, like collagen, gelatin, chitosan, alginate, hyaluronic acid (HA), and silk fibroin, have become promising scaffolds because they are bioactive, mimic the extracellular matrix (ECM), and can be broken down by enzymes. Crosslinking and composite reinforcement can greatly change how well they work. For example, collagen scaffolds that are highly crosslinked with glutaraldehyde keep up to 51.9% of their tensile strength after being exposed to enzymes, while non-crosslinked scaffolds only keep 12% of their strength. Chitosan–hydroxyapatite matrices, on the other hand, can reach compressive strengths of 2–12 MPa, which is close to the strength of cancellous bone. Additive manufacturing and 4D printing allow for precise control of structures and the ability to change their shape over time, which helps with vascularization and mechanical adaptation. Injectable and in situ-forming hydrogels show clinically important results, such as filling 85% of osteochondral defects in rabbits, improving left ventricular ejection fraction by up to 9% in large-animal cardiac models, and speeding up healing by 25–40% in chronic wounds. Even with these improvements, it is still hard to get batch consistency, a standardized way to test mechanical properties, and production that meets GMP (Good Manufacturing Practices) standards and can be scaled up. Full article

Brucellosis is a globally prevalent zoonosis that causes abortion and infertility in livestock, leading to substantial economic losses. Sensitive and reliable quantification of Brucella antibodies, particularly at trace levels, is critical for early diagnosis. In this work, an electrochemical immunosensor was developed by integrating electric field-assisted antigen immobilization with an electrode platform. The electrode was first electrochemically pretreated to improve interfacial reproducibility, and then sequentially modified with L-cysteine and glutaraldehyde to construct an antigen-coupling layer. During antigen immobilization, a custom-built electric field device was applied to regulate the interfacial arrangement of Brucella antigens. The fabrication process was characterized by scanning electron microscopy and cyclic voltammetry, and the analytical performance was evaluated by electrochemical impedance spectroscopy and voltammetric measurements. Under the optimized conditions, the proposed immunosensor exhibited a linear response to Brucella antibodies over the range of 1 × 10⁻⁶–10 IU/mL, with a correlation coefficient of 0.99 and a detection limit of 2.04 × 10⁻⁷ IU/mL. The sensor also showed acceptable specificity, repeatability, and short-term storage stability, with recoveries of 93.15–99.14% in spiked milk samples. These results indicate that electric field-assisted immobilization can serve as a useful interfacial regulation strategy for Brucella immunosensing and support the analytical feasibility of the proposed platform under controlled experimental conditions. Further validation in more complex biological matrices is still required. Full article

Chitosan-coated sand has been developed as a sustainable, environmentally friendly, and cost-effective water treatment method for removing nitrate anions, leveraging the adsorption properties of chitosan. When applied to sand using glutaraldehyde as a cross-linking agent, this adsorbent removes nitrate anions with an adsorption capacity (q_e) of 154.41 mg g⁻¹. This approach is particularly advantageous due to its low cost, high adsorption capacity, and effectiveness over a wide range of pH and temperatures, although its performance is optimal under slightly acidic to neutral conditions (pH = 6) due to electrostatic attraction and ion exchange, as the positively charged amino groups of chitosan bind to the negatively charged nitrate ions. Nitrate adsorption is also described by the Langmuir isotherm and follows the pseudo-second-order model. Furthermore, the adsorbent remains highly stable even after five regeneration cycles, demonstrating its long-term effectiveness and durability, while offering a cost-effective and environmentally friendly solution in accordance with the principles of sustainable development. Full article

Homeobox protein MEIS2 has been strongly implicated in colorectal cancer (CRC) progression and metastatic potential, making its targeted inhibition a promising therapeutic strategy. However, recently developed MEIS inhibitors are limited by poor aqueous solubility, instability under physiological conditions, and insufficient intracellular accumulation, which restrict their clinical applicability. To overcome these challenges, a multifunctional hybrid nanoemulsome system was developed by integrating boron–silane-doped carbon dots (CDs) with chitosan via glutaraldehyde crosslinking, followed by emulsification with oleic acid and non-ionic surfactants (Span 80 and Tween 20/80) in the presence of a MEIS inhibitor (MEISi-2). The resulting composite exhibited high structural stability, excellent biocompatibility, and a drug encapsulation efficiency of 96.2%. Fourier-transform infrared spectroscopy (FTIR) and dynamic light scattering (DLS) analyses confirmed successful hybridization and the formation of nanoemulsions with an average particle size of approximately 320 nm following drug loading. The system demonstrated controlled drug release under physiological conditions. In vitro studies using HCT116 CRC and HaCaT healthy keratinocytes revealed effective cellular uptake and selective cytotoxicity. The intrinsic fluorescence properties of CDs enabled real-time monitoring of intracellular drug delivery via DAPI-channel imaging. Overall, this hybrid nanoemulsome platform provides a stable and efficient delivery system for MEIS inhibitors and represents a promising strategy for the treatment of CRC. Furthermore, this approach may be extended to other poorly soluble amphiphilic therapeutic agents. Full article

To enhance the functionality of collagen (Coll)-based scaffolds, we developed a double-crosslinking strategy incorporating an electroconductive chitosan (Ch) and polypyrrole (Ppy) composite. Successful pre-crosslinking of Ch and Ppy was achieved using glutaraldehyde (GTA) at 100 µM. This facilitated imine linkage formation, confirmed by FTIR, enabling synergistic integration with Coll and successful nanofiber scaffold fabrication via electrospinning. While increasing the Ch-Ppy-GTA ratio affected the spinning process and higher GTA concentrations compromised fiber homogeneity, all other measured properties generally improved with increasing ratios. Crucially, this methodology allowed the membranes to maintain their morphology and significantly extended their degradation profile up to 20–30 days in PBS medium at 37 °C. Furthermore, the scaffolds exhibited electroactivity characterized by pseudocapacitance in the presence of Na⁺ and Ca²⁺ ions. These findings demonstrate a robust, tunable method for creating electroactive and structurally stable nanofiber scaffolds suitable for advanced tissue engineering. Full article

Background and Clinical Significance: To our knowledge, there is a lack of electron microscopy studies in hematocornea since 1985, and more so for graft hematocornea after deep anterior lamellar keratoplasty (DALK). This study provides an ultrastructural characterization of hematocornea occurring in a DALK graft. Our study presents several limitations: single-case design and lack of control tissue. Case Presentation: The DALK graft with hematocornea was excised and introduced inside of the operating room in glutaraldehyde solution recipient. The graft was quickly cold-transported to light and transmission electron microscopy. Hematocornea in the DALK transplant graft resulted in features of stromal alteration and dysfunctional cellular clean-up response. The collagen lamellae ultrastructure was affected near electron-dense hem deposits. Two cellular aspects were observed: adaptation and degeneration. Electron-dense granules were found in keratocytes, which may exhibit cellular adaptations, such as vacuoles and phagosomes. Macropinocytosis may mechanistically explain ingestion of electron-dense granules, and dysfunctions in the macropinocytosis process may have led to cell degeneration. Cellular degeneration was marked by loss of organelle contour and loss of cellular membrane integrity (burst-cell aspect). Microscopic corneal alteration corresponded to macroscopic total loss of corneal transparency and elasticity. Conclusions: This study described lamellar ultrastructure alterations and dysfunctional cellular response in hematocornea of a DALK corneal transplant graft. Full article

Antimicrobial resistance (AMR) has become a major global public health challenge, and widely residual disinfectants in the environment are one of the key drivers of bacterial AMR development. This study aimed to investigate the inductive effects of three commonly used disinfectants—benzalkonium bromide (BAB), glutaraldehyde (GTA), and povidone-iodine (PVP-I)—on the resistance of Escherichia coli (E. coli), as well as the resultant bacterial phenotypic and genetic alterations. Three disinfectants frequently detected in clinical and environmental settings were selected as the research objects: first, their bactericidal efficacy against environmental bacteria was determined; subsequently, a concentration-increasing gradient approach was adopted to systematically explore the evolutionary patterns of E. coli resistance under the stress of sub-inhibitory concentrations (SICs). After induction, the bacterial resistance levels to disinfectants and various antibiotics, growth characteristics, and biofilm-forming ability were detected, and combined with whole-genome analysis to investigate genetic-level changes. The results showed that all three disinfectants could enhance E. coli resistance to themselves (12–48-fold) and antibiotics, and the induced antibiotic resistance exhibited favorable genetic stability. Among them, BAB induced the strongest resistance, with the most significant increase in resistance levels to multiple antibiotics (16–64-fold); GTA had the weakest inductive effect, only slightly enhancing bacterial resistance to a small number of antibiotics. Notably, all induced strains exhibited reduced growth rates yet markedly enhanced biofilm-forming capacity, alongside acquired genomic structural variations. Their gene functions displayed shared adaptive signatures in coping with environmental stress, while core pathogenicity-associated genes remained conserved. This study demonstrates that inducing E. coli using environmentally relevant low concentrations of disinfectant residues as initial induction doses drives the evolution of bacterial antimicrobial resistance (AMR), with distinct resistance induction risks among the three disinfectant types. These findings offer critical insights for standardizing disinfectant application, mitigating the transmission of bacterial AMR, and underscore the imperative of interdisciplinary collaboration to tackle the environmental risks posed by disinfectant residues. Full article