Diagnostic value of tuberculosis-specific antigens ESAT-6 and CFP10 in lymph node tuberculosis.

Objective: To assess the diagnostic value of immunohistochemical (IHC) staining for detecting the tuberculosis-secreted antigens ESAT-6 and CFP10 in lymph node tuberculosis. Methods: Archived, paraffin-embedded lymph node specimens from 72 patients diagnosed with lymph node tuberculosis and 68 patients with lymphoma were retrospectively collected from the Department of Pathology at the Affiliated Hospital of North Sichuan Medical College, Nanchong, Sichuan Province, China between January 2016 and March 2023. These specimens were subjected to acid-fast and immunohistochemical staining to compare the effectiveness of these methods, with their sensitivity and specificity evaluated against a comprehensive reference standard. Results: Acid-fast staining demonstrated a sensitivity of 12.3% and a specificity of 100%. IHC staining for ESAT-6 showed a sensitivity of 87.5% and a specificity of 85.3%, whereas IHC staining for CFP10 exhibited a sensitivity of 75.0% and a specificity of 89.7%. Conclusion: The study indicates that IHC detection of ESAT-6 and CFP10 in paraffin-embedded lymph node tuberculosis tissues has a markedly higher sensitivity compared to acid-fast staining. Thus, IHC staining may serve as a supplementary diagnostic tool for the pathological evaluation of lymph node tuberculosis.

Ultrastiff metamaterials generated through a multilayer strategy and topology optimization.

Metamaterials composed of different geometrical primitives have different properties. Corresponding to the fundamental geometrical forms of line, plane, and surface, beam-, plate-, and shell-based lattice metamaterials enjoy many advantages in many aspects, respectively. To fully exploit the advantages of each structural archetype, we propose a multilayer strategy and topology optimization technique to design lattice metamaterial in this study. Under the frame of the multilayer strategy, the design space is enlarged and diversified, and the design freedom is increased. Topology optimization is applied to explore better designs in the larger and diverse design space. Beam-plate-shell-combined metamaterials automatically emerge from the optimization to achieve ultrahigh stiffness. Benefiting from high stiffness, energy absorption performances of optimized results also demonstrate substantial improvements under large geometrical deformation. The multilayer strategy and topology optimization can also bring a series of tunable dimensions for lattice design, which helps achieve desired mechanical properties, such as isotropic elasticity and functionally grading material property, and superior performances in acoustic tuning, electrostatic shielding, and fluid field tuning. We envision that a broad array of synthetic and composite metamaterials with unprecedented performance can be designed with the multilayer strategy and topology optimization.

Insecticidal potential of a Consolida ajacis extract and its major compound (ethyl linoleate) against the diamondback moth, Plutella xylostella.

The diamondback moth (Plutella xylostella) is one of the most destructive lepidopteran pests of cruciferous vegetables. However, DBM has developed resistance to current chemical and biological insecticides used for its control, indicating the necessity for finding new insecticides against it. Bio-insecticides derived from plant extracts are eco-friendly alternatives to synthetic pesticides. The aims of this study were to evaluate the insecticidal activity of Consolida ajacis seed extracts against DBM, the underlying mechanism of the control effect of promising extracts, and the identification of the main insecticidal compounds of these extracts. The results showed that ethyl acetate extract of C. ajacis seed exhibited strong contact toxicity (LC50: 5.05 mg/mL), ingestion toxicity, antifeedant, and oviposition deterrent activities against DBM, among the extracts evaluated. At 72 h, glutathiase, acetylcholinesterase, carboxylesterase, peroxidase, and superoxide dismutase activities were inhibited, but catalase activity was activated. The main compound identified from the extract was ethyl linoleate, which had the most significant insecticidal activity on the diamondback moths. This study's findings provide a better understanding of the insecticidal activity of ethyl acetate extract obtained from C. ajacis and its main component (ethyl linoleate). This will help in the development of new insecticides to control DBM.

Electrochemical fluorescence switching of enhanced green fluorescent protein.

Switchable fluorescent proteins, for which fluorescence can be switched ON and OFF, are widely used for molecule tracking and super resolution imaging. However, the robust use of the switchable fluorescent proteins is still limited as either the switching is not repeatable, or such switching requires irradiation with coupled lasers of different wavelengths. Herein, we report an electrochemical approach to reversible fluorescence switching for enhanced green fluorescent proteins (EGFP) on indium tin oxide coated glass. Our results demonstrate that negative and positive electrochemical potentials can efficiently switch the fluorescent proteins between the dim (OFF) and bright (ON) states at the single molecule level. The electrochemical fluorescence switching is fast, reversible, and may be performed up to hundreds of cycles before photobleaching occurs. These findings highlight that this method of electrochemical fluorescence switching can be incorporated into advanced fluorescence microscopy.

Machine Learning Color Feature Analysis of a High Throughput Nanoparticle Conjugate Sensing Assay.

Plasmonic nanoparticles are finding applications within the single molecule sensing field in a "dimer" format, where interaction of the target with hairpin DNA causes a decrease in the interparticle distance, leading to a localized surface plasmon resonance shift. While this shift may be detected using spectroscopy, achieving statistical relevance requires the measurement of thousands of nanoparticle dimers and the timescales required for spectroscopic analysis are incompatible with point-of-care devices. However, using dark-field imaging of the dimer structures, simultaneous digital analysis of the plasmonic resonance shift after target interaction of thousands of dimer structures may be achieved in minutes. The main challenge of this digital analysis on the single-molecule scale was the occurrence of false signals caused by non-specifically bound clusters of nanoparticles. This effect may be reduced by digitally separating dimers from other nanoconjugate types. Variation in image intensity was observed to have a discernible impact on the color analysis of the nanoconjugate constructs and thus the accuracy of the digital separation. Color spaces wherein intensity may be uncoupled from the color information (hue, saturation, and value (HSV) and luminance, a* vector, and b* vector (LAB) were contrasted to a color space which cannot uncouple intensity (RGB) to train a classifier algorithm. Each classifier algorithm was validated to determine which color space produced the most accurate digital separation of the nanoconjugate types. The LAB-based learning classifier demonstrated the highest accuracy for digitally separating nanoparticles. Using this classifier, nanoparticle conjugates were monitored for their plasmonic color shift after interaction with a synthetic RNA target, resulting in a platform with a highly accurate yes/no response with a true positive rate of 88% and a true negative rate of 100%. The sensor response of tested single stranded RNA (ssRNA) samples was well above control responses for target concentrations in the range of 10 aM-1 pM.

Aggregation-induced emission-active micelles: synthesis, characterization, and applications.

Aggregation-induced emission (AIE)-active micelles are a type of fluorescent functional materials that exhibit enhanced emissions in the aggregated surfactant state. They have received significant interest due to their excellent fluorescence efficiency in the aggregated state, remarkable processability, and solubility. AIE-active micelles can be designed through the self-assembly of amphipathic AIE luminogens (AIEgens) and the encapsulation of non-emissive amphipathic molecules in AIEgens. Currently, a wide range of AIE-active micelles have been constructed, with a significant increase in research interest in this area. A series of advanced techniques has been used to characterize AIE-active micelles, such as cryogenic-electron microscopy (Cryo-EM) and confocal laser scanning microscopy (CLSM). This review provides an overview of the synthesis, characterization, and applications of AIE-active micelles, especially their applications in cell and in vivo imaging, biological and organic compound sensors, anticancer drugs, gene delivery, chemotherapy, photodynamic therapy, and photocatalytic reactions, with a focus on the most recent developments. Based on the synergistic effect of micelles and AIE, it is anticipated that this review will guide the development of innovative and fascinating AIE-active micelle materials with exciting architectures and functions in the future.

Liver-Targeted Near-Infrared Fluorescence/Photoacoustic Dual-Modal Probe for Real-Time Imaging of In Situ Hepatic Inflammation.

Early diagnosis of hepatic inflammation is the key to timely treatment and avoid the worsening of liver inflammation. Near-infrared fluorescence (NIRF) probes have high sensitivity but low spatial resolution in lesion imaging, while photoacoustic (PA) imaging has good spatial location information. Therefore, the development of a NIRF/PA dual-modal probe integrated with high sensitivity and spatial location feedback can achieve an accurate early diagnosis of hepatic inflammation. Here, we report an activatable NIRF/PA dual-modal probe (hCy-Tf-CA) for the detection of the superoxide anion (O2·-) in early hepatic inflammation. hCy-Tf-CA showed high selectivity and sensitivity for detecting O2·- fluctuation in vitro. More importantly, by introducing hepatocyte-targeting cholic acid (CA), the probe successfully achieved accurate in situ imaging of acute inflammatory liver injury (AILI) and autoimmune hepatitis (AIH) in vivo. The introduced CA not only promotes the hepatic targeting accumulation of probes but also improves the performance of low background dual-modal imaging in vivo. Therefore, hCy-Tf-CA provides an effective strategy for significantly improving in situ imaging performance and holds great potential for early, sensitive, and accurate diagnosis of hepatic inflammation.

Statistical predictions on the encapsulation of single molecule binding pairs into sized-dispersed nanocontainers.

Single molecule experiments have recently attracted enormous interest. Many of these studies involve the encapsulation of a single molecule into nanoscale containers (such as vesicles, droplets and nanowells). In such cases, the single molecule encapsulation efficiency is a key parameter to consider in order to get a statistically significant quantitative information. It has been shown that such encapsulation typically follows a Poisson distribution and such theory of encapsulation has only been applied to the encapsulation of single molecules into perfectly sized monodispersed containers. However, experimentally nanocontainers are usually characterized by a size distribution, and often just a single binding pair (rather than a single molecule) is required to be encapsulated. Here the use of Poisson distribution is extended to predict the encapsulation efficiency of two different molecules in an association equilibrium. The Poisson distribution is coupled with a log-normal distribution in order to consider the effect of the container size distribution, and the effect of adsorption to the container is also considered. This theory will allow experimentalists to determine what single molecule encapsulation efficiency can be expected as a function of the experimental conditions. Two case studies, based on experimental data, are given to support the theoretical predictions.

Nanoparticle-based single molecule fluorescent probes.

Single molecule fluorescent probes have attracted considerable attention due to their ultimate sensitivity, fast response, low sample consumption, and high signal-to-noise ratio. Nanoparticles with outstanding optical properties make them perfect candidates for probes in the application of single molecule detection. In this review, we focus on various kinds of nanoparticles acting as single molecule fluorescent probes, including quantum dots, upconverting fluorescent nanoparticles, carbon dots, single-wall carbon nanotubes, fluorescent nanodiamonds, polymeric nanoparticles, nanoclusters, and metallic nanoparticles. Optical properties of various nanoparticles and their recent application in single molecule fluorescent probes are explored. How nanoparticles boost the sensitivity of detection is emphasized in combination with different sensing strategies. Future trends of nanoparticles in single molecule detection are also discussed. We hope that this review can provide practical guidance for researchers who work on nanoparticle-based single molecule fluorescent probes.

Chemiluminescent Probes Based on 1,2-Dioxetane Structures For Bioimaging.

Chemiluminescent probes based on 1,2-dioxetane scaffold are one of the most sensitive imaging modalities for detecting disease-related biomarkers and can obtain more accurate biological information in cells and in vivo. Due to the elimination of external light excitation, the background autofluorescence problem in fluorescence technology can be effectively avoided, providing ultrahigh sensitivity and signal-to-noise ratio for various applications. In this review, we highlight a comprehensive but concise overview of activatable 1,2-dioxetane-based chemiluminescent probes by reporting significant advances in accurate detection and bioimaging. The design principles and applications for reactive species, enzymes, and other disease-related biomarkers are systematically discussed and summarized. The challenges and potential prospects of chemiluminescent probes are also discussed to further promote the development of new chemiluminescence methods for biological analysis and diagnosis.

Enzyme-activated near-infrared fluorogenic probe with high-efficiency intrahepatic targeting ability for visualization of drug-induced liver injury.

Hepatotoxicity is a serious problem faced by thousands of clinical drugs, and drug-induced liver injury (DILI) caused by chronic administration or overdose has become a major biosafety issue. However, the near-infrared (NIR) fluorescent probes currently used for liver injury detection still suffer from poor liver targeting ability and low sensitivity. Enzyme-activated fluorogenic probes with powerful in situ targeting ability are the key to improving the imaging effect of liver injury. Herein, we rationally designed a leucine aminopeptidase (LAP) activated fluorogenic probe hCy-CA-LAP, which greatly improved the hepatocyte-targeting capability by introducing a cholic acid group. The probe hCy-CA-LAP is converted into a high-emission hCy-CA fluorophore in the presence of LAP, showing high selectivity, high sensitivity and low detection limit (0.0067 U mL-1) for LAP, and successfully realizes the sensitive detection of small fluctuations of LAP in living cells. Moreover, the probe can achieve effective in situ accumulation in the liver, thereby achieving precise imaging and evaluation of two different types of drug-induced hepatotoxicity in vivo. Therefore, the probe hCy-CA-LAP may be a potential tool for exploring the roles of LAP and evaluating the degree of DILI.

Bioorthogonal chemistry in metal clusters: a general strategy for the construction of multifunctional probes for bioimaging in living cells and in vivo.

Multifunctional bioimaging probes based on metal clusters have multiple characteristics of metal clusters and functional conjugates, and their development has broad application prospects in the fields of biomedical imaging and tumor diagnosis. However, current bioconjugation methods on metal clusters are time-consuming and have low reaction efficiency, which hinders the construction of bioimaging probes with multifunctional components. Here, we report a concise and promising design strategy to realize the simple and efficient introduction of functional conjugates through bioorthogonal reactions based on azido-functionalized metal clusters. Based on this strategy, taking the probe FA-CuC@BSA-Cy5 as an example, we demonstrated the design of a copper cluster-based multifunctional near-infrared (NIR) fluorescent probe and its real-time imaging application in vivo. Through the strain-promoted azide-alkyne cycloaddition (SPAAC) reaction, the tumor-specific targeting ligand folic acid (FA) and fluorophore (Cy5) can be chemically conjugated to azido-functionalized CuC@BSA-N3 quickly and efficiently under biocompatible conditions. The prepared probe showed numerous advantages of metal clusters, including good stability, ultra-small particle size and low toxicity and rapid renal clearance. At the same time, FA-modified FA-CuC@BSA-Cy5 can specifically target KB cells with high FR expression, and in vivo fluorescence imaging shows higher tumor accumulation. The construction of the azido functional metal cluster platform can be extended to various metal clusters with functional probes and prodrugs, thereby providing more promising candidates for future medical diagnoses.

Surface modification by poly(ethylene glycol) with different end-grafted groups: Experimental and theoretical study.

Dihydroxyphenylalanine (DOPA) is extensively reported to be a surface-independent anchor molecule in bioadhesive surface modification and antifouling biomaterial fabrication. However, the mechanisms of DOPA adsorption on versatile substrates and the comparison between experimental results and theoretical results are less addressed. We report the adsorption of DOPA anchored monomethoxy poly(ethylene glycol) (DOPA-mPEG) on substrates and surface wettability as well as antifouling property in comparison with thiol and hydroxyl anchored mPEG (mPEG-SH and mPEG-OH). Gold and hydroxylated silicon were used as model substrates to study the adsorptions of mPEGs. The experimental results showed that the DOPA-mPEG showed higher affinity to both gold and silicon wafers, and the DOPA-mPEG modified surfaces had higher resistance to protein adsorption than those of mPEG-SH and mPEG-OH. It is revealed that the surface wettability is primary for surface fouling, while polymer flexibility is the secondary parameter. We present ab initio calculations of the adsorption of mEGs with different end-functionalities on Au and hydroxylated silicon wafer (Si-OH), where the binding energies are obtained. It is established that monomethoxy ethylene glycol (mEG) with DOPA terminal DOPA-mEG is clearly favored for the adsorption with both gold and Si-OH surfaces due to the bidentate Au-O interactions and the bidentate O-H bond interactions, in agreement with experimental evidence.

Genome-wide identification and characterization of WOX genes in Cucumis sativus.

WUSCHEL-related homeobox (WOX) proteins are plant-specific transcription factors that are profoundly involved in regulation of plant development and stress responses. In this study, we totally identified 11 WOX transcription factor family members in cucumber (Cucumis sativus, CsWOX) genome and classified them into three clades with nine subclades based on phylogenetic analysis results. Alignment of amino acid sequences revealed that all WOX members in cucumber contained the typical homeodomain, which consists of 60-66 amino acids and is folded into a helix-turn-helix structure. Gene duplication event analysis indicated that CsWOX1a and CsWOX1b were a segment duplication pair, which might affect the number of WOX members in cucumber genome. The expression profiles of CsWOX genes in different tissues demonstrated that the members sorted into the ancient clade (CsWOX13a and CsWOX13b) were constitutively expressed at higher levels in comparison to the others. Cis-element analysis in promoter regions suggested that the expression of CsWOX genes was associated with phytohormone pathways and stress responses, which was further supported by RNA-seq data. Taken together, our results provide new insights into the evolution of cucumber WOX genes and improve our understanding about the biological functions of the CsWOX gene family.

Impact of the Coverage of Aptamers on a Nanoparticle on the Binding Equilibrium and Kinetics between Aptamer and Protein.

Knowledge of the interaction between aptamer and protein is integral to the design and development of aptamer-based biosensors. Nanoparticles functionalized with aptamers are commonly used in these kinds of sensors. As such, studies into how the number of aptamers on the nanoparticle surface influence both kinetics and thermodynamics of the binding interaction are required. In this study, aptamers specific for interferon gamma (IFN-γ) were immobilized on the surface of gold nanoparticles (AuNPs), and the effect of surface coverage of aptamer on the binding interaction with its target was investigated using fluorescence spectroscopy. The number of aptamers were adjusted from an average of 9.6 to 258 per particle. The binding isotherm between AuNPs-aptamer conjugate and protein was modeled with the Hill-Langmuir equation, and the determined equilibrium dissociation constant (K'D) decreased 10-fold when increasing the coverage of aptamer. The kinetics of the reaction as a function of coverage of aptamer were also investigated, including the association rate constant (kon) and the dissociation rate constant (koff). The AuNPs-aptamer conjugate with 258 aptamers per particle had the highest kon, while the koff was similar for AuNPs-aptamer conjugates with different surface coverages. Therefore, the surface coverage of aptamers on AuNPs affects both the thermodynamics and the kinetics of the binding. The AuNPs-aptamer conjugate with the highest surface coverage is the most favorable in biosensors considering the limit of detection, sensitivity, and response time of the assay. These findings deepen our understanding of the interaction between aptamer and target protein on the particle surface, which is important to both improve the scientific design and increase the application of aptamer-nanoparticle based biosensor.

Activatable Off-on Near-Infrared QCy7-based Fluorogenic Probes for Bioimaging.

The activatable off-on near-infrared QCy7-based fluorogenic probes have emerged as powerful modalities for detecting and monitoring biological analytes and understanding their biological processes in cells and organisms. The use of biomarker-activated QCy7-based probes enables simple synthesis, minimum photo-damage to biological samples, and minimum background interference from biological systems. In this minireview, we aim to provide a rigorous but concise overview of activatable QCy7-based fluorogenic probes by reporting the significant progress made in recent years. The design strategies and the main applications of accurate detection and imaging of disease-related biomarkers (including ROS/RSS, enzymes, metal ions, and other related species) were reasonably analyzed and discussed. The potential challenges and prospects of activatable QCy7-based fluorogenic probes are also emphasized to further advance the development of new methods for biomarker detection and bioimaging.

Genome-wide analysis of the WRKY gene family in the cucumber genome and transcriptome-wide identification of WRKY transcription factors that respond to biotic and abiotic stresses.

BACKGROUND: Cucumber (Cucumis sativus L.) is an economically important vegetable crop species. However, it is susceptible to various abiotic and biotic stresses. WRKY transcription factors play important roles in plant growth and development, particularly in the plant response to biotic and abiotic stresses. However, little is known about the expression pattern of WRKY genes under different stresses in cucumber. RESULTS: In the present study, an analysis of the new assembly of the cucumber genome (v3.0) allowed the identification of 61 cucumber WRKY genes. Phylogenetic and synteny analyses were performed using related species to investigate the evolution of the cucumber WRKY genes. The 61 CsWRKYs were classified into three main groups, within which the gene structure and motif compositions were conserved. Tissue expression profiles of the WRKY genes demonstrated that 24 CsWRKY genes showed constitutive expression (FPKM > 1 in all samples), and some WRKY genes showed organ-specific expression, suggesting that these WRKYs might be important for plant growth and organ development in cucumber. Importantly, analysis of the CsWRKY gene expression patterns revealed that five CsWRKY genes strongly responded to both salt and heat stresses, 12 genes were observed to be expressed in response to infection from downy mildew and powdery mildew, and three CsWRKY genes simultaneously responded to all treatments analysed. Some CsWRKY genes were observed to be induced/repressed at different times after abiotic or biotic stress treatment, demonstrating that cucumber WRKY genes might play different roles during different stress responses and that their expression patterns vary in response to stresses. CONCLUSIONS: Sixty-one WRKY genes were identified in cucumber, and insight into their classification, evolution, and expression patterns was gained in this study. Responses to different abiotic and biotic stresses in cucumber were also investigated. Our results provide a better understanding of the function of CsWRKY genes in improving abiotic and biotic stress resistance in cucumber.

A novel amplified electrochemiluminescence biosensor based on Au NPs@PDA@CuInZnS QDs nanocomposites for ultrasensitive detection of p53 gene.

In this work, a novel surface plasmon resonance (SPR) enhanced electrochemiluminescence (ECL) biosensing model was first designed based on Au NPs@polydopamine (PDA)@CuInZnS QDs nanocomposite. Au NPs were coated with the PDA layer via the electrostatic force. CuInZnS QDs were bound on the surface of Au NPs@PDA nanocomposite. CuInZnS QDs worked as ECL luminophore in the sensing application. PDA shell not only controlled the separation length between Au NPs and QDs to induce SPR enhanced ECL response, but also limited the potential charge transfer and ECL quenching effect. As a result, the nanocomposite ECL intensity was twice that of QDs with K2S2O8. The tumor suppressor p53 gene was detected in the amplified ECL sensing system. The sensing method has a linear response in the range of 0.1 nmol/L to 15 nmol/L with a detection limit of 0.03 nmol/L. The DNA biosensor based on the nanocomposite showed excellent sensitivity, selectivity, reproducibility and stability and was applied in spiked human serum samples with satisfactory results.

Ultrasensitive detection of EGFR gene based on surface plasmon resonance enhanced electrochemiluminescence of CuZnInS quantum dots.

In our work, a novel DNA electrochemiluminescence (ECL) sensor based on CuZnInS quantum dots (QDs) and gold-nanoparticles (Au NPs) is developed for highly sensitive detection of epidermal growth factor receptor (EGFR) Gene, which has a close relation with the lung cancer. The CuZnInS QDs work as a novel kind of ECL luminophore, whose defect state emission is suitable for ECL sensing. To enhance the sensitivity of the sensing system, Au NPs are utilized creatively to strengthen the ECL intensity of CuZnInS QDS according to the surface plasmon resonance (SPR) effect. An ultrasensitive and universal detecting platform is built based on the SPR effect between Au NPs and CuZnInS QDS. The effect of the capped stabilizer on the ECL signal of QDs is firstly investigated. Three different stabilizers are used to cap the CuZnInS QDs, including mercaptopropionic acid (MPA), l-glutathione (GSH) and cysteamine (CA). MPA capped CuZnInS QDs possess the strongest ECL intensity among the three kinds of the CuZnInS QDs. Under the optimum conditions, a good linear relationship between ECL intensity and the concentration of target DNA is obtained in the range from 0.05 nmol L-1 to 1 nmol L-1. The detection limit is 0.0043 nmol L-1. The proposed DNA sensor has been employed for the determination of target DNA EGFR in human serum samples with satisfactory results.