Search Results (1,380)

Protein-based APIs represent a big group of modern therapeutics. Their characterization involves complex analytical protocols which require special methods, especially in the case when the protein drug is included into tablet dosage forms. Although the fluorescence polarization assay (FPA) is not currently regulated by many national Pharmacopeias, it represents a promising approach for protein drug standardization, considering their rapid, sensitive, and automatable detection suitable for high-throughput analysis and real-time quality control. To evaluate the applicability of FPA for the analysis of protein drugs in tablets, the quantifying of lysozyme in tablet dosage forms was studied by this method with the use of a fluorescently labeled synthetic chitooligosaccharide tracer. It was shown that this approach overcomes the limitations of the conventional turbidimetric assay of lysozyme determination, which is labor-intensive and relies on unstable reagents. Measurements were performed with both portable and stationary fluorescence polarization readers. Commercial tablets from five manufacturers containing lysozyme (20 mg) and pyridoxine hydrochloride (10 mg) together with other excipients were analyzed. The FPIA method showed a linear range of 5.0–70 µg/mL, with specificity confirmed by the absence of interference from excipients. Accuracy, evaluated by standard addition (10–20 mg), yielded recoveries of 100.2–106.0%. Placebo spiked with lysozyme at 80–120% of nominal content demonstrated recoveries of 98.0–100.1%, with RSD (n = 6) not exceeding 13.7%, indicating good precision. The developed method enables reliable lysozyme quantification in tablets, offering speed, simplicity, and robustness, and shows its suitability for the routine quality control of protein-containing dosage forms including the enzyme ones. Full article

Background/Objectives: The cell surface proteoglycan glypican-3 (GPC3) is reportedly overexpressed in hepatocellular carcinoma (HCC) tissues, but not in benign liver tissues, rendering this protein a potential target for radionuclide theranostic approaches. Peptides are generally a promising class of targeting molecules for the development of radioligands because they combine straightforward synthetic access with favorable pharmacokinetics. Among the published peptides with disclosed structures, one of the most promising radioligands is [¹⁸F]AlF-NOTA-TJ12P2, which has a reported comparably high binding affinity to GPC3 and a high hydrophilicity. In this study, we aimed to design novel GPC3-targeting radioligands based on the TJ12P2 peptidic scaffold. Methods: Peptides were synthesized on solid phase using an Fmoc protecting group strategy. For comparative investigations, the reference nanobody HN3 was expressed in E. coli, isolated and subsequently modified with NODA-GA or SulfoCy3. The binding of native peptides, scrambled variants and reference nanobodies to GPC3 was investigated by surface plasmon resonance (SPR) interaction analysis, and fluorescently labeled versions of peptides and nanobodies were used for fluorescence microscopy in HepG2 (GPC3+) or SK Hep1 (GPC3−) cells. The chelator-bearing peptides were radiolabeled with gallium-67 and their stability towards radiolysis and in human serum was investigated. The binding of radiolabeled peptides and nanobodies to HepG2 cells was assessed in real-time ligand binding experiments. Results: The synthesized native peptides did not exhibit binding towards GPC3 in SPR interaction analyses, and the observed response was comparable to that of the scrambled variants at equal concentrations. Additionally, no binding to or uptake of the fluorescent constructs into cells was observed with fluorescence microscopy regardless of cellular GPC3 expression level. In real-time radioligand binding experiments, very fast association and dissociation of the gallium-67 labeled peptides to GPC3 positive HepG2 cells was observed, suggesting either extremely fast binding kinetics or unspecific binding of the peptides. Conclusions: Taken together, these findings suggest that the peptide TJ12P2 lacks specific binding to GPC3 in vitro and might not serve as a basis for the development of radioligands targeting GPC3. Full article

Salicylic acid (SA) is a key phytohormone that coordinates plant innate immunity and systemic acquired resistance. Because SA levels and signaling are highly dynamic in space and time, a suite of SA-focused tools, including SA-specific microbial biosensors and SA-responsive transcriptional and chemical reporters, has been developed to study them. This review compares three classes of tools in terms of sensitivity, specificity, temporal resolution, invasiveness, quantifiability, and suitability across species. We describe developing genetically encoded sensors that can directly sense salicylic acid and report it, for example, via a fluorescence resonance energy transfer signal or another real-time output. We offer recommendations on method selection by research goal and plant species, as well as combined protocols (long-term autoluminescence plus local probes/biosensors) for cross-validation. Future work should prioritize substrate-free, quantitative SA reporters deployable in crops and the field; coupled with CRISPR-based editing and screening, these tools would enable reporter-guided discovery of immunity genes and rapid engineering of durable disease resistance. Full article

YABBY belongs to the family of plant-specific transcription factors, known for their role in plant morphology, growth, and development. Its name is derived from the first discovered member—the YABBY1 gene of Arabidopsis thaliana (named due to its mutated phenotype showing a “Y-shaped” bifurcation). Despite extensive research across various plant species, no studies have conducted a genome-wide investigation of the YABBY gene family in Cerasus humilis. This study identified six ChYABBY (Cerasus humilis YABBY) genes distributed across five chromosomes through a comprehensive bioinformatic analysis of the C. humilis genome. The gene expression during the four growth phases was confirmed using real-time-quantitative fluorescent PCR (qPCR). ChYABBY is segmented into five distinct subfamilies. Genetic lineage analysis determined the close genetic relationship between the YABBY genes of C. humilis and Malus pumila. An examination of the gene architecture and preserved motifs revealed that ChYABBY typically comprises 5–6 introns, with motif1, motif2, and motif3 being preserved domains across all ChYABBY protein sequences. Promoter analysis suggests that ChYABBY genes play various roles in the growth and maturation of C. humilis. An examination of the homology revealed the absence of tandem replication in the ChYABBY gene family, with a single pair of fragment-replicating genes. The heat map and q-PCR results indicate that the expression of the ChYABBY gene is tissue-specific and correlates with some aspects of the fruit growth and development. This suggests a potential role for this gene family in fruit maturation. The determination of total sugar and total flavonoid content indicated that the content of the two substances was high when the fruit was green. The antioxidant capacity of the fruit at each stage was different. This research provides an important basis for further understanding the structure and function of the ChYABBY gene, and lays a foundation for the identification of YABBY genes in Rosaceae plants. Full article

Optical fiber technology is becoming essential in modern radiation therapy, enabling precise, real-time, and minimally invasive monitoring. As oncology moves toward patient-specific treatment, there is growing demand for adaptable and biologically compatible sensing tools. Fiber-optic systems meet this need by integrating into clinical workflows with highly localized dosimetric and spectroscopic feedback. Their small size and flexibility allow deployment within catheters, endoscopes, or treatment applicators, making them suitable for both external beam and internal therapies. This paper reviews the fundamental principles and diverse applications of optical fiber sensing technologies in radiation oncology, focusing on dosimetry, spectroscopy, imaging, and adaptive radiotherapy. Implementations such as scintillating and Bragg grating-based dosimeters demonstrate feasibility for in vivo dose monitoring. Spectroscopic techniques, such as Raman and fluorescence spectroscopy, offer real-time insights into tissue biochemistry, aiding in treatment response assessment and tumor characterization. However, despite such advantages of optical fiber sensors, challenges such as signal attenuation, calibration demands, and limited dynamic range remain. This paper further explores clinical application, technical limitations, and future directions, emphasizing multiplexing capabilities, integration and regulatory considerations, and trends in machine learning development. Collectively, these optical fiber sensing technologies show strong potential to improve the safety, accuracy, and adaptability of radiation therapy in personalized cancer care. Full article

ZnO nanostructures have attracted attention as transducer materials in optical biosensing platforms due to their wide bandgap, defect-mediated photoluminescence, high surface-to-volume ratio, and tunable morphology. This review examines how the dimensionality of ZnO nanostructures affects biosensor performance, particularly in terms of charge transport, signal transduction, and biomolecule immobilization. The synthesis approaches are discussed, highlighting how they influence crystallinity, defect density, and surface functionalization potential. The impact of immobilization strategies on sensor stability and sensitivity is also assessed. The role of ZnO in various optical detection schemes, including photoluminescence, surface plasmon resonance (SPR), localized (LSPR), fluorescence, and surface-enhanced Raman scattering (SERS), is reviewed, with emphasis on label-free and real-time detection. Representative case studies demonstrate the detection of clinically and environmentally relevant targets, such as glucose, dopamine, cancer biomarkers, and SARS-CoV-2 antigens, with limits of detection in the pico- to femtomolar range. Recent developments in ZnO-based hybrid systems and their integration into fiber-optic and microfluidic platforms are explored as scalable solutions for portable, multiplexed diagnostics. The review concludes by outlining current challenges related to reproducibility, long-term operational stability, and surface modification standardization. This work provides a framework for understanding structure–function relationships in ZnO-based biosensors and highlights future directions for their development in biomedical and environmental monitoring applications. Full article

Tuberculosis remains a serious global public health challenge and requires the development of rapid, sensitive, and specific diagnostic tools for effective treatment and disease control. Bioimaging reporters are promising diagnostic tools that exploit the unique biochemical properties of Mycobacterium tuberculosis for real-time detection of viable cells from clinical samples. Moreover, these methods offer significant advantages over the conventional methods currently used in practice, including reduced assay time, increased specificity, and the ability to discriminate viable cells from dead cells. In this review, we highlight reporters of a different nature that the enable direct detection of Mycobacterium tuberculosis, eliminating complex sample preparation. Such reporters could serve as powerful tools in fluorescence microscopy, provide alternative strategies for automated culture-based diagnostic systems, and offer new approaches for developing point-of-care methods and diagnostic devices suitable for clinical practice. Full article

Cellular uptake of nanomaterials is based on endocytosis with their endosomal–lysosomal entrapment resulting in enzymatic hydrolysis. Besides biodegradation, the antigen presentation induces innate and adaptive immunity. Our goal was isolation of extracellular particles to study their structures, penetration into cells, stability, intracellular distribution, and interferon (IFN) production. Extracellular nanomaterials were isolated from conditioned culture media of human embryonic and cancer cells by two-stage differential centrifugation. Cellular uptake of Cy5-labeled particles was evaluated using spectrofluorimetry and confocal fluorescent microscopy. IFN gene expression was analyzed by reverse transcription with real-time PCR and ELISA. Vesicles of 10–200 nm were isolated by centrifugation at 20,800× g at +4 °C for 30 min. The fluorescent vesicles were gradually accumulated inside cells for seven days. Intracellular distribution patterns of the Cy5-labeled vesicles differed from lysosomes stained with LysoRed tracker. IFNs α, β and γ were not detected after treatment with the vesicles. IFN λ was found in cells in the presence of allogenic but not autologous particles. The gradual cellular uptake occurred without significant differences between autologous and heterologous vesicles. Different localization of the extracellular vesicles (EV) and lysosomes along with weak innate immune response (if any) suggested membrane fusion. Full article

Stray cats (Felis vaga) are key hosts for feline and zoonotic pathogens. From June to August 2024, we conducted a cross-sectional study across six districts in Shenzhen, China, involving 126 cats sampled from three types of sites. Multiple specimens were tested via quantitative real-time PCR (qPCR) for feline coronavirus type I (FCoV-I), feline calicivirus (FCV), feline herpesvirus type I (FHV-I), feline panleukopenia virus (FPV), and rabies virus (RABV); serum was analyzed for RABV-neutralizing antibodies by the fluorescent antibody virus neutralization (FAVN) assay. The overall pathogen positivity was 89.68%. FPV was most prevalent (61.90%), followed by FCV (57.14%), FCoV-I (46.83%), and FHV-I (23.02%). No RABV nucleic acid was detected. The co-infection rate reached 62.70%, primarily dual infections (33.33%). Geographical variation was observed, with significantly higher FCoV-I in Longgang than Futian (p < 0.05). RABV seropositivity was only 6.00%. FCV and FPV co-occurred most frequently (Jaccard = 0.456). All pathogen pairs had relative risk (RR) > 1, suggesting non-random co-infections, though not significant after correction. In summary, major feline pathogens are widespread with frequent co-infections among Shenzhen stray cats, while low rabies immunity indicates potential public health risk. Targeted control measures are warranted. Full article

Background/Objectives: Lyme disease diagnosis remains challenging due to the limitations of current methods. While PCR-based assays are widely used, their sensitivity can be affected by sample type and the inhibition of host DNA. This study aimed to evaluate the feasibility and sensitivity of a CRISPR/Cas12-based detection system for Borrelia burgdorferi sensu lato, comparing its performance with real-time PCR. Methods: DNA from three Borrelia genospecies (B. burgdorferi, B. garinii, and B. afzelii) was amplified targeting the OspA gene. Detection was performed using a Cas12/crRNA system with a fluorescent ssDNA reporter. Sensitivity assays were conducted on serial dilutions of Borrelia DNA, with and without human genomic DNA, and results were compared with qPCR. Results: Direct detection of Borrelia DNA without amplification was not feasible. However, when combined with PCR, the Cas12/crRNA system reliably detected as few as 5 genome copies per reaction. End-point PCR extended to 60 cycles improved detection robustness for B. garinii and B. afzelii, although sensitivity decreased in the presence of human genomic DNA. Conclusions: The Cas12/crRNA-based system offers a sensitive and accessible alternative to qPCR, especially in settings lacking real-time PCR instrumentation. Future developments may include integration with isothermal amplification and microfluidic platforms to enhance direct detection capabilities. Full article

Early detection and diagnosis of plant diseases is critical for ensuring global food security and sustainable agricultural practices. This review comprehensively examines latest advancements in crop disease risk prediction, onset detection through imaging techniques, machine learning (ML), deep learning (DL), and edge computing technologies. Traditional disease detection methods, which rely on visual inspections, are time-consuming, and often inaccurate. While chemical analyses are accurate, they can be time consuming and leave less flexibility to promptly implement remedial actions. In contrast, modern techniques such as hyperspectral and multispectral imaging, thermal imaging, and fluorescence imaging, among others can provide non-invasive and highly accurate solutions for identifying plant diseases at early stages. The integration of ML and DL models, including convolutional neural networks (CNNs) and transfer learning, has significantly improved disease classification and severity assessment. Furthermore, edge computing and the Internet of Things (IoT) facilitate real-time disease monitoring by processing and communicating data directly in/from the field, reducing latency and reliance on in-house as well as centralized cloud computing. Despite these advancements, challenges remain in terms of multimodal dataset standardization, integration of individual technologies of sensing, data processing, communication, and decision-making to provide a complete end-to-end solution for practical implementations. In addition, robustness of such technologies in varying field conditions, and affordability has also not been reviewed. To this end, this review paper focuses on broad areas of sensing, computing, and communication systems to outline the transformative potential of end-to-end solutions for effective implementations towards crop disease management in modern agricultural systems. Foundation of this review also highlights critical potential for integrating AI-driven disease detection and predictive models capable of analyzing multimodal data of environmental factors such as temperature and humidity, as well as visible-range and thermal imagery information for early disease diagnosis and timely management. Future research should focus on developing autonomous end-to-end disease monitoring systems that incorporate these technologies, fostering comprehensive precision agriculture and sustainable crop production. Full article

This study reports the construction and validation of a CAFLUX (Chemically Activated Fluorescent Expression) HepG2 reporter cell line engineered to express a histone H2B–green fluorescent protein (H2B–GFP) fusion protein under the control of a dioxin-responsive cytochrome P450 1A1 (CYP1A1) promoter. A lentiviral construct containing a synthetic promoter with multiple dioxin-responsive elements (DREs) upstream of the H2B–EGFP coding sequence was cloned into the pFUGW vector, packaged in human embryonic kidney (HEK) 293FT cells, and used to transduce HepG2 hepatocellular carcinoma cells. Stable clones obtained by limiting dilution were screened for GFP expression in response to 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD). The resulting CAFLUX HepG2 cells exhibited dose-dependent nuclear GFP fluorescence when exposed to aryl hydrocarbon receptor (AhR) agonists, with limits of detection of approximately 0.01 pM for TCDD and 0.1 pM for benzo[a]pyrene (B[a]P), a polycyclic aromatic hydrocarbon (PAH). This reporter activity correlated with endogenous CYP1A1 mRNA expression as determined by quantitative polymerase chain reaction (qPCR), confirming that GFP signals reflected native transcriptional responses. In functional assays, curcumin suppressed GFP expression in a concentration-dependent manner and induced apoptotic morphology at higher doses, while extracellular vesicles (EVs) derived from adipose-derived stem cells (ADSCs) significantly reduced both GFP fluorescence and CYP1A1 mRNA levels, suggesting an inhibitory effect on AhR-driven transcription. The CAFLUX HepG2 reporter system therefore provides a sensitive and reproducible platform for real-time, nuclear-localized monitoring of AhR-mediated gene expression. Its responsiveness to both agonists and antagonists underscores its potential utility in toxicological evaluation, drug discovery, and the investigation of EV-mediated signaling in liver cancer models. Full article

Single-molecule detection (SMD) has reformed analytical science by enabling the direct observation of individual molecular events, thus overcoming the limitations of ensemble-averaged measurements. This review presents a comprehensive analysis of the principles, devices, and emerging materials that have shaped the current landscape of SMD. We explore a wide range of sensing mechanisms, including surface plasmon resonance, mechanochemical transduction, transistor-based sensing, optical microfiber platforms, fluorescence-based techniques, Raman scattering, and recognition tunneling, which offer distinct advantages in terms of label-free operation, ultrasensitivity, and real-time responsiveness. Each technique is critically examined through representative case studies, revealing how innovations in device architecture and signal amplification strategies have collectively pushed the detection limits into the femtomolar to attomolar range. Beyond the sensing principles, this review highlights the transformative role of advanced nanomaterials such as graphene, carbon nanotubes, quantum dots, MnO₂ nanosheets, upconversion nanocrystals, and magnetic nanoparticles. These materials enable new transduction pathways and augment the signal strength, specificity, and integration into compact and wearable biosensing platforms. We also detail the multifaceted applications of SMD across biomedical diagnostics, environmental monitoring, food safety, neuroscience, materials science, and quantum technologies, underscoring its relevance to global health, safety, and sustainability. Despite significant progress, the field faces several critical challenges, including signal reproducibility, biocompatibility, fabrication scalability, and data interpretation complexity. To address these barriers, we propose future research directions involving multimodal transduction, AI-assisted signal analytics, surface passivation techniques, and modular system design for field-deployable diagnostics. By providing a cross-disciplinary synthesis of device physics, materials science, and real-world applications, this review offers a comprehensive roadmap for the next generation of SMD technologies, poised to impact both fundamental research and translational healthcare. Full article

Sulfur dioxide (SO₂) is commonly employed as an antioxidant and preservative in food processing, but excessive intake of SO₂ can pose significant health risks. Therefore, accurate detection of sulfite content in food is crucial for ensuring food quality and safety. A novel fluorescent probe, BODIPY-Y, composed of a BODIPY derivative and an ethyl cyanoacetate group linked by a carbon–carbon double bond, was synthesized for detecting sulfur dioxide derivatives. When the BODIPY-Y probe interacts with SO₃²⁻, the probe exhibits enhanced fluorescence at 514 nm. Spectrometric experiments show that the probe exhibits high sensitivity (LOD: 0.263 μmol/L), a fast response time (50 s) and excellent selectivity for SO₃²⁻. Mechanistic studies confirm that the BODIPY-Y probe operates via an intramolecular charge transfer (ICT) mechanism. The carbon–carbon double bond in BODIPY-Y undergoes nucleophilic addition with SO₃²⁻, blocking the ICT process and resulting in a blue shift in the fluorescence spectrum. In addition, the probe was applied to quantify SO₃²⁻ levels in real food samples. The measured concentrations of SO₂ in the white sugar and red wine were 15.93 μmol/L and 7.30 μmol/L, respectively, with recovery rates of 77.9–98.1%. This work presents a prospective chemical tool for monitoring sulfur dioxide derivatives in food products. Full article

The increasing prevalence of antimicrobial resistance and the persistent challenge of infectious diseases highlight the critical necessity for novel approaches that integrate pathogen management with swift detection methods. Carbon dots (CDs) are a versatile class of fluorescent nanomaterials that have garnered increasing attention owing to their tunable surface chemistry, strong photoluminescence, high stability, and biocompatibility. Recent studies demonstrate that CDs possess broad-spectrum antibacterial and antifungal activities via multiple mechanisms, including the generation of reactive oxygen species, disruption of membranes, inhibition of biofilms, and synergistic interactions with conventional antimicrobials. The performance is significantly affected by precursor selection, heteroatom doping, and surface functionalization, with minimum inhibitory concentrations reported to range from highly potent at the microgram level to moderate at elevated concentrations. The intrinsic fluorescence of CDs, in addition to their antimicrobial activity, facilitates their use as sensitive and selective probes for microbial detection, allowing for rapid and real-time monitoring in biomedical, food safety, and environmental settings. This review summarizes recent advancements in the antimicrobial properties of carbon dots (CDs) and their fluorescence-based applications in microbial detection. It emphasizes their theranostic potential and future prospects as multifunctional nanomaterials for combating infectious diseases and ensuring microbial safety. Full article