Design of an Antigen-Triggered Nanobody-Based Fluorescence Probe for PET Immunoassay to Detect Quinalphos in Food Samples.

Photoinduced electron-transfer (PET) immunoassay based on a fluorescence site-specifically labeled nanobody, also called mini Quenchbody (Q-body), exhibits extraordinary sensitivity and saves much time in the homogeneous noncompetitive mode and is therefore regarded as a valuable method. However, limited by the efficiency of both quenching and dequenching of the fluorescence signal before and after antigen binding associated with the PET principle, not all original nanobodies can be used as candidates for mini Q-bodies. Herein, with the anti-quinalphos nanobody 11A (Nb-11A) as the model, we, for the first time, adopt a strategy by combining X-ray structural analysis with site-directed mutagenesis to design and produce a mutant Nb-R29W, and then successfully generate a mini Q-body by labeling with ATTO520 fluorescein. Based on this, a novel PET immunoassay is established, which exhibits a limit of detection of 0.007 µg/mL with a detection time of only 15 min, 25-fold improved sensitivity, and faster by 5-fold compared to the competitive immunoassay. Meanwhile, the recovery test of vegetable samples and validation by the standard ultraperformance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) both demonstrated that the established PET immunoassay is a novel, sensitive, and accurate detection method for quinalphos. Ultimately, the findings of this work will provide valuable insights into the development of triggered PET fluorescence probes by using existing antibody resources.

A Rotation-Inversion Dual-Motion Molecular Switch: Race for NMR Signaling.

The interplay between the thermal helical inversion (THI) of the stiff-stilbene moiety and the rotation of the dimethylamino (DMA) group in 1 results in a dependence of the DMA NMR signals on the THI kinetics in (E)-1 but the rotation kinetics in (Z)-1, because the faster motion mode is responsible. Consequently, the photochemical switching from (E)-1 to (Z)-1 illustrates the phenomenon of "switchable motion detection" by the same set of NMR signals in a dual-motion molecular system.

Novel Polycyclic Fused Amide Derivatives: Properties and Applications for Sky-blue Electroluminescent Devices.

Aromatic imide derivatives play a critical role in boosting the electroluminescent (EL) performance of organic light-emitting diodes (OLEDs). However, the majority of aromatic imide-based materials are limited to long wavelength emission OLEDs rather than blue emissions due to their strong electron-withdrawing characteristics. Herein, two novel polycyclic fused amide units were reported as electron acceptor to be combined with either a tetramethylcarbazole or acridine donor via a phenyl linker to generate four conventional fluorescence blue emitters of BBI-4MeCz, BBI-DMAC, BSQ-4MeCz and BSQ-DMAC for the first time. BSQ-4MeCz and BSQ-DMAC based on a BSQ unit exhibited higher thermal stability and photoluminescence quantum yields than BBI-4MeCz and BBI-DMAC based on a BBI unit due to their more planar acceptor structure. The intermolecular interactions that exist in the BSQ series materials effectively inhibit the molecular rotation and configuration relaxation, and thus allow for blue-shifted emissions. Blue OLED devices were constructed with the developed materials as emitters, and the effects of both the structure of the polycyclic fused amide acceptor and the electron donor on the EL performance were clarified. Consequently, a sky-blue OLED device based on BSQ-DMAC was created, with a high maximum external quantum efficiency of 4.94% and a maximum luminance of 7761 cd m-2.

The Most Active Oxidase-Mimicking Mn2 O3 Nanozyme for Biosensor Signal Generation.

Oxidase-mimicking nanozymes are more desirable than peroxidase-mimicking ones since H2 O2 can be omitted. However, only a few nanomaterials are known for oxidase-like activities. In this work, we compared the activity of Mn2 O3 , Mn3 O4 and MnO2 and found that Mn2 O3 had the highest oxidase activity. Interestingly, the activity of Mn2 O3 was even inhibited by H2 O2 . The oxidase-like activity of Mn2 O3 was not much affected by the presence of proteins such as bovine serum albumin (BSA), but the physisorption of antibodies to Mn2 O3 was not strong enough to withstand the displacement by BSA. We then treated Mn2 O3 with 3-aminopropyltriethoxysilane to graft an amine group, which was used to conjugate antibodies using glutaraldehyde as a crosslinker. A one-step indirect competitive ELISA (icELISA) was developed for the detection of isocarbophos, and an IC50 of 261.7 ng/mL was obtained, comparable with the results of the standard two-step assay using horseradish peroxidase (HRP)-labeled antibodies. This assay has the advantage of significant timesaving for rapid detection of large amounts of samples. This work has discovered a highly efficient oxidase-mimicking nanozyme useful for various nano- and analytical applications.

Effect of proteins on the oxidase-like activity of CeO2 nanozymes for immunoassays.

Having the benefits of low cost, excellent stability and tolerance to extreme conditions, nanozymes are a potential alternative of horseradish peroxidase (HRP) or other enzymes for bioanalytical chemistry, especially immunoassays. CeO2 nanoparticles have oxidase-mimicking activity and can avoid the use of unstable H2O2. For robust assays, the effect of proteins on the activity of CeO2 needs to be carefully studied. Herein, we studied the adsorption and desorption of bovine serum albumin (BSA) from CeO2. The CeO2 nanoparticles exhibited a higher protein adsorption capacity compared to the other tested metal oxide nanoparticles. Although the oxidase-like activity of CeO2 was inhibited by BSA, low concentrations of phosphate and fluoride ions boosted the activity of protein-capped CeO2. CeO2 was still active under strong acidic conditions and at high temperature, while HRP lost its activity. For immunoassay development, we covered CeO2 with an amine-modified silane for covalent conjugation to antibodies. A one-step indirect competitive ELISA for fenitrothion was developed, and an IC50 value of 35.6 ng mL-1 and a limit of detection of 2.1 ng mL-1 were obtained.

Predictive value of cadherin-11 for subsequent recurrence and progression in non-muscle invasive bladder cancer.

BACKGROUND: Cadherin-11 (CDH11) is a type II cadherin and reported to function as an oncogene in various cancers. Our present study aims to investigate the role of CDH11 in bladder cancer (BCA). METHODS: Bioinformatics analysis was performed in four independent microarray data including 56 non-muscle-invasive bladder cancer (NMIBC) and 132 muscle-invasive bladder cancer (MIBC) tissues from Gene Expression Omnibus to screen out differentially expressed genes. Next, we detected CDH11 expression in BCA specimens and cell lines by qPCR and western blotting assays. Immunohistochemical analyses were performed in 209 paraffin-embedded BCA samples and 30 adjacent normal bladder tissues. RESULTS: Bioinformatics analysis revealed that CDH11 had a higher expression level in MIBC tissues than in NMIBC, which was consistent with our clinical BCA specimens and cell lines at both mRNA and protein levels. Immunohistochemical analysis demonstrated that over-expression of CDH11 was closely related to the histological grade, pT status, tumour size and poor outcomes of BCA patients. What's more, CDH11 (area under curve (AUC) = 0.673 and 0.735) had a better predictive value than E-cadherin (AUC = 0.629 and 0.629) and a similar discrimination with the European Organization for Research and Treatment of Cancer (EORTC) score system (AUC = 0.719 and 0.667) in evaluating potential recurrence and progression of NMIBC. Moreover, combination of CDH11 and EORTC score system was the best predictive model in predicting recurrence of NMIBC (AUC = 0.779) among the three models. CONCLUSIONS: CDH11 was a reliable therapeutic target in BCA and a useful index to predict the possibilities of recurrence and progression in NMIBC patients.

Production of a specific monoclonal antibody for 1-naphthol based on novel hapten strategy and development of an easy-to-use ELISA in urine samples.

1-naphthol (1-NAP) is the main metabolite of pesticide carbaryl and naphthalene, and is also a genotoxic and carcinogenic intermediate in the synthesis of organic compound, dyes, pigment and pharmaceutical industry. In this work, two novel haptens were designed and synthesized for developing a competitive indirect enzyme-linked immunosorbent assay (ciELISA) method for 1-NAP in urine samples. The assay showed a limit of detection of 2.21 ng/mL and working range from 4.02 ng/mL to 31.25 ng/mL for 1-NAP in optimized working buffer. The matrix effect of samples was eliminated via 15-fold dilution of optimized working buffer. Good average recoveries (102.4%-123.4%) with a coefficient of variation from 11.7% to 14.7% was obtained for spiked urine samples. Subsequent instrument verification test showed good correlation between the results of ciELISA and high-performance liquid chromatography. The developed ciELISA is a high-throughput tool to monitor 1-NAP in urine, which can provide technical support for the establishment of biological exposure level for the exposure to carbaryl, naphthalene and other related pollutants.

Production of Antigen-Binding Fragment against O,O-Diethyl Organophosphorus Pesticides and Molecular Dynamics Simulations of Antibody Recognition.

Immunoassay for pesticides is an emerging analytical method since it is rapid, efficient, sensitive, and inexpensive. In this study, a recombinant antigen-binding fragment (Fab) against a broad set of O,O-diethyl organophosphorus pesticides (DOPs) was produced and characterized. The κ chain and Fd fragment were amplified via PCR and inserted into the vector pComb3XSS and the soluble Fab on phagemid pComb3XSS was induced by isopropyl β-d-thiogalactoside in E. coli TOP 10F’. SDS-PAGE, Western blotting, and indirect competitive ELISA results indicated that Fab maintained the good characteristics of the parental mAb. To better understand antibody recognition, the three-dimensional (3D) model of Fab was built via homologous modeling and the interaction between Fab and DOPs was studied via molecular docking and dynamics simulations. The model clearly explained the interaction manner of Fab and DOPs, and showed that the Arg-L96 and Arg-H52 were mainly responsible for antibody binding. This work provided a foundation for further mutagenesis of Fab to improve its characteristics.

The expression of cysteine-rich secretory protein 2 (CRISP2) and its specific regulator miR-27b in the spermatozoa of patients with asthenozoospermia.

Cysteine-rich secretory protein 2 (CRISP2) is an important sperm protein and plays roles in spermatogenesis, modulation of flagellar motility, acrosome reaction, and gamete fusion. Clinical evidence shows a reduced CRISP2 expression in spermatozoa from asthenozoospermic patients, but the molecular mechanism underlying its reduction remains unknown. Herein, we carried out a study focusing on the CRISP2 reduction and its roles in asthenozoospermia. Initially, through analyzing CRISP2 expression and methylation on CRISP2 promoter activity in sperm, we observed a decreased expression of CRISP2 protein rather than its mRNA in the ejaculated spermatozoa from asthenozoospermic patients and no methylation in the CRISP2 promoter, suggesting CRISP2 expression may be regulated in the sperm at the posttranscriptional level. Subsequently, we found that microRNA 27b (miR-27b), predicted as a candidate regulator of CRISP2 using bioinformatics, was highly expressed in the ejaculated spermatozoa from asthenozoospermic patients. Luciferase reporter assay and transfection experiments disclosed that this microRNA could target CRISP2 by specifically binding its 3' untranslated region, suppressing CRISP2 expression. Extended clinical observation further confirmed a highly expressed miR-27b and its obviously negative correlation with CRISP2 protein expression in ejaculated spermatozoa samples from asthenozoospermic patients. Finally, we conducted a retrospective follow-up study to support that either high miR-27b expression or low CRISP2 protein expression was significantly associated with low sperm progressive motility, abnormal morphology, and infertility. Thus, this study provides the first preliminary insight into the mechanism leading to the reduced CRISP2 expression in asthenozoospermia, offering a potential therapeutic target for treating male infertility or for male contraception.

GAA/(Au-Au/IrO2)@Cu(PABA) reactor with cascade catalytic activity for α-glucosidase inhibitor screening.

BACKGROUND: In vitro screening strategies based on the inhibition of α-glucosidase (GAA) activity have been widely used for the discovery of potential antidiabetic drugs, but they still face some challenges, such as poor enzyme stability, non-reusability and narrow range of applicability. To overcome these limitations, an in vitro screening method based on GAA@GOx@Cu-MOF reactor was developed in our previous study. However, the method was still not satisfactory enough in terms of construction cost, pH stability, organic solvent resistance and reusability. Thence, there is still a great need for the development of in vitro screening methods with lower cost and wider applicability. RESULTS: A colorimetric sensing strategy based on GAA/(Au-Au/IrO2)@Cu(PABA) cascade catalytic reactor, which constructed through simultaneous encapsulating Au-Au/IrO2 nanozyme with glucose oxidase-mimicking and peroxidase-mimicking activities and GAA in Cu(PABA) carrier with peroxidase-mimicking activity, was innovatively developed for in vitro screening of GAA inhibitors in this work. It was found that the reactor not only exhibited excellent thermal stability, pH stability, organic solvent resistance, room temperature storage stability, and reusability, but also possessed cascade catalytic performance, with approximately 12.36-fold increased catalytic activity compared to the free system (GAA + Au-Au/IrO2). Moreover, the in vitro GAA inhibitors screening method based on this reactor demonstrated considerable anti-interference performance and detection sensitivity, with a detection limit of 4.79 nM for acarbose. Meanwhile, the method owned good reliability and accuracy, and has been successfully applied to the in vitro screening of oleanolic acid derivatives as potential GAA inhibitors. SIGNIFICANCE: This method not only more effectively solved the shortcomings of poor stability, narrow scope of application, and non-reusability of natural enzymes in the classical method compared with our previous work, but also broaden the application scope of Au-Au/IrO2 nanozyme with glucose oxidase and peroxidase mimicking activities, and Cu(PABA) carrier with peroxidase mimicking activity, which was expected to be a new generation candidate method for GAA inhibitor screening.

Simple dual-readout immunosensor based on phosphate-triggered and potassium permanganate for visual detection of fenitrothion.

Multicolor-based visual immunosensor is a promising tool for rapid analysis without the use of bulky instruments. Herein, an anti-fenitrothion nanobody-alkaline phosphatase fusion protein (VHHjd8-ALP) was employed to develop a multicolor visual immunosensor (MVIS) and a ratiometric fluorescence MVIS (RFMVIS, respectively). After one-step competitive immunoassay, the VHHjd8-ALP bound to microplate catalyzed phenyl phosphate disodium salt (ArP) into phenol. Under high alkaline condition (pH 12), the phenol reduced KMnO4 to intermediate (K2MnO4) and further to MnO2 in alkaline condition (pH 12), accompanied by a visible color transition of purple-green-yellow, which can be used for semiquantitative visual analysis or qualitative detection by measuring RGB value. RFMVIS was proposed on the basis of MVIS to further improve sensitivity. The CdTe quantum dot and fluorescein were used as signal probes to develop the fluorescent immunosensor. The CdTe dots with red emission (644 nm) was quenched by oxidation of KMnO4, whereas the fluorescein with green emission (520 nm) remained constant, accompanied by a fluorescent color transition of green-yellow-red. By measuring the ratio of the fluorescence intensity (I644/I520), the ratiometric fluorescence immunosensor was developed for qualitative analysis. The two visual immunosensors were sensitive and simple, and they showed good accuracy and practicability in the recovery test, thus are ideal tools for rapid screening.

Development of broad-spectrum immunoassay with monoclonal antibody to detect five eugenols and study of their molecular recognition mechanism.

In this study, three eugenol fragment-containing haptens were synthesized, and a monoclonal antibody (mAb) selective for five commonly-found eugenol compounds (EUGs, i.e., eugenol, isoeugenol, methyl eugenol, methyl isoeugenol, and acetyl isoeugenol) was obtained. Based on this mAb, a broad-spectrum indirect competitive ELISA for high-throughput detection of five EUGs was developed. The detection limits for eugenol, isoeugenol, methyl eugenol, methyl isoeugenol and acetyl isoeugenol in both tilapia and shrimp samples were 25.3/ 50.6 µg/kg, 0.075/0.15 µg/kg, 0.48/0.96 µg/kg, 0.16/0.32 µg/kg, and 18.16/36.32 µg/kg, respectively. The recoveries for five EUGs ranged from 80.4 to 114.0 % with a coefficient of variation less than 11.5 %. Moreover, homology modelling and molecular docking were conducted to elucidate the interactions mechanism of mAb-EUGs. The work provides a promising tool for high-throughput screening of EUGs in aquatic products, which can serve as a benchmark for designing haptens and developing immunoassays for other small molecules.

Scalable Fabrication of Polymeric Composite Microspheres to Inhibit Oral Pathogens and Promote Osteogenic Differentiation of Periodontal Membrane Stem Cells.

Periodontitis is a worldwide bacterial infectious disease, resulting in the resorption of tooth-supporting structures. Biodegradable polymeric microspheres are emerging as an appealing local therapy candidate for periodontal defect regeneration but suffer from tedious procedures and low yields. Herein, we developed a facile yet scalable approach to prepare polylactide composite microspheres with outstanding drug-loading capability. It was realized by blending equimolar polylactide enantiomers at the temperature between the melting point of homocrystallites and stereocomplex (sc) crystallites, enabling the precipitation of sc crystallites in the form of microspheres. Meanwhile, epigallocatechin gallate (EGCG) and nano-hydroxyapatite were encapsulated in the microspheres in the designated amount. Such an assembly allowed the fast and sustained release of EGCG and Ca2+ ions. The resultant hybrid composite microspheres not only exhibited strong antimicrobial activity against typical oral pathogens (Porphyromonas gingivalis and Enterococcus faecalis), but also directly promoted osteogenic differentiation of periodontal ligament stem cells with good cytocompatibility. These dual-functional composite microspheres offer a desired drug delivery platform to address the practical needs for periodontitis treatment.

A bispecific nanobody with high sensitivity/efficiency for simultaneous determination of carbaryl and its metabolite 1-naphthol in the soil and rice samples.

The simultaneous determination of carbaryl and its metabolite 1-naphthol is essential for risk assessment of pesticide exposure in agricultural and environmental samples. Herein, several bispecific nanobodies (BsNbs) with different lengths of hydrophilic linkers and junction sites were prepared and characterized for the simultaneous recognition of carbaryl and its metabolite 1-naphthol. It was found that the affinity of BsNbs to the analytes could be regulated by controlling linker length and linking terminal. Additionally, molecular simulation revealed that linker lengths affected the conformation of BsNbs, leading to alteration in sensitivity. The BsNb with G4S linker, named G4S-C-N-VHH, showing good thermal stability and sensitivity was used to develop a bispecific indirect competitive enzyme-linked immunosorbent assay (Bic-ELISA). The assay demonstrated a limit of detection of 0.8 ng/mL for carbaryl and 0.4 ng/mL for 1-naphthol in buffer system. Good recoveries from soil and rice samples were obtained, ranging from 80.0% to 112.7% (carbaryl) and 76.5%-110.8% (1-naphthol), respectively. Taken together, this study firstly provided a BsNb with high sensitivity and efficiency against environmental pesticide and its metabolite, and firstly used molecular dynamics simulation to explore the influence of linker on recognition. The results are valuable for the application of immunoassay with high efficiency in the fields of environment and agriculture.

Dual-modular immunosensor for bongkrekic acid detection using specific monoclonal antibody.

Bongkrekic acid (BA) is a mitochondrial toxin that causes high mortality but is often mistakenly categorized as other food poisonings. The immunoassay of BA is still challenging since the specific antibody is unavailable. In this work, a monoclonal antibody specific to BA was first generated and a dual-modular immunosensor for on-site and laboratory detection was established. The antibody showed good affinity (Kd=0.33 µM) and sensitivity (IC50 =17.9 ng/mL in ELISA) with negligible cross-reactivity with common mycotoxins. In dual-modular conditions, fluorescence assay (FA) was conducted based on the inner filter effect of carbon dots (CDs) and oxidized 3,3',5,5'-tetramethylbenzidine (TMB), while the colorimetric assay (CA) was conducted using TMB2+-mediated rapid surface etching of gold nanostars (Au NSs). The proposed immunosensor showed good sensitivity and reproducibility to BA in food samples, with a limit of detection lower than 10 ng/mL and recovery ranging from 80.0% to 103.6%, which was in good consistence with that of standard LC-MS/MS. Overall, the proposed immunosensor is an ideal tool for screening BA contaminants in food with good sensitivity and high effectivity.

Development of a Biotinylated Nanobody-Based Gold Nanoparticle Immunochromatographic Assay for the Detection of Procymidone in Crops.

A heavy-chain antibody (VHH) library against procymidone (PRM) was constructed via immunizing Bactrian camels. Through careful biopanning, seven nanobodies (Nbs) with different sequences were obtained. The variability in their performance was primarily attributed to the amino acid differences in complementarity-determining region 3 (CDR3), as analyzed by molecular docking. The Nb exhibiting the highest sensitivity, named NbFM5, was biotinylated and conjugated to streptavidin-labeled gold nanoparticles to preserve the epitope's activity and prevent a decrease in sensitivity due to traditional random electrostatic adsorption. Subsequently, a simple and sensitive immunochromatographic assay (ICA) was developed for rapid detection of PRM based on biotinylated Nb (btNb). The developed btNb-ICA showed a cut-off value of 200 ng/mL for visual judgment and a half-inhibitory concentration (IC50) of 6.04 ng/mL for quantitative detection. The limit of detection (LOD) was as low as 0.88 ng/mL. The recoveries in actual samples of crops ranged from 82.2 to 117.3%, aligning well with the results obtained from GC-MS/MS (R2 = 0.995). In summary, the developed btNb-ICA demonstrated high specificity and good accuracy for the rapid detection of PRM residues in vegetables. The total analysis time from preparing the sample to obtaining the result was less than 25 min.

Modeling Dynamic Recrystallization Behavior in a Novel HIPed P/M Superalloy during High-Temperature Deformation.

The dynamic recrystallization (DRX) features and the evolution of the microstructure of a new hot isostatic pressed (HIPed) powder metallurgy (P/M) superalloy are investigated by hot-compression tests. The sensitivity of grain dimension and DRX behavior to deformation parameters is analyzed. The results reveal that the DRX features and grain-growth behavior are significantly affected by deformation conditions. The DRX process is promoted with a raised temperature/true strain or a reduced strain rate. However, the grains grow up rapidly at relatively high temperatures. At strain rates of o.1 s-1 and 1 s-1, a uniform microstructure and small grains are obtained. Due to the obvious differences in the DRX rate at various temperatures, the piecewise DRX kinetics equations are proposed to predict the DRX behavior. At the same time, a mathematical model for predicting the grain dimension and the grain growth behavior is established. To further analyze the DRX behavior and the changes in grain dimension, the hot deformation process is simulated. The developed grain-growth equation as well as the piecewise DRX kinetics equations are integrated into DEFORM software. The simulated DRX features are consistent with the test results, indicating that the proposed DRX kinetics equations and the established grain-growth model can be well used for describing the microstructure evolution. So, they are very useful for the practical hot forming of P/M superalloy parts.

Rapid and Wash-Free Time-Gated FRET Histamine Assays Using Antibodies and Aptamers.

Histamine (HA) is an indicator of food freshness and quality. However, high concentrations of HA can cause food poisoning. Simple, rapid, sensitive, and specific quantification can enable efficient screening of HA in food and beverages. However, conventional assays are complicated and time-consuming, as they require multiple incubation, washing, and separation steps. Here, we demonstrate that time-gated Förster resonance energy transfer (TG-FRET) between terbium (Tb) complexes and organic dyes can be implemented in both immunosensors and aptasensors for simple HA quantification using a rapid, single-step, mix-and-measure assay format. Both biosensors could quantify HA at concentrations relevant in food poisoning with limits of detection of 0.19 µg/mL and 0.03 µg/mL, respectively. Excellent specificity was documented against the structurally similar food components tryptamine and l-histidine. Direct applicability of the TG-FRET assays was demonstrated by quantifying HA in spiked fish and wine samples with both excellent concentration recovery and agreement with conventional multistep enzyme-linked immunosorbent assays (ELISAs). Our results show that the simplicity and rapidity of TG-FRET assays do not compromise sensitivity, specificity, and reliability, and both immunosensors and aptasensors have a strong potential for their implementation in advanced food safety screening.

Polycaprolactone Electrospun Nanofiber Membrane with Sustained Chlorohexidine Release Capability against Oral Pathogens.

Multiple-pathogen periodontal disease necessitates a local release and concentration of antibacterial medication to control inflammation in a particular location of the mouth cavity. Therefore, it is necessary to effectively load and deliver medicine/antibiotics to treat numerous complex bacterial infections. This study developed chlorhexidine (CHX)/polycaprolactone (PCL) nanofiber membranes with controlled release properties as periodontal dressings to prevent or treat oral disorders. Electrostatic spinning was adopted to endow the nanofiber membranes with a high porosity, hydrophilicity, and CHX loading capability. The release of CHX occurred in a concentration-dependent manner. The CHX/PCL nanofiber membranes exhibited good biocompatibility with human periodontal ligament stem cells, with cell viability over 85% in each group via CCK-8 assay and LIVE/DEAD staining; moreover, the good attachment of the membrane was illustrated by scanning electron microscopy imaging. Through the agar diffusion assay, the nanofiber membranes with only 0.075 wt% CHX exhibited high antibacterial activity against three typical oral infection-causing bacteria: Porphyromonas gingivalis, Enterococcus faecalis, and Prevotella intermedia. The results indicated that the CHX/PCL nanofiber holds great potential as a periodontal dressing for the prevention and treatment periodontal disorders associated with bacteria.

Phosphate-triggered ratiometric fluoroimmunoassay based on nanobody-alkaline phosphatase fusion for sensitive detection of 1-naphthol for the exposure assessment of pesticide carbaryl.

The excessive use of carbaryl has resulted in the risk of its exposure. In this study, we isolated six nanobodies (Nbs) from a camelid phage display library against the biomarker of carbaryl, 1-naphthol (1-NAP). Owing to its characteristics of easy genetic modifications, we produced a nanobody-alkaline phosphatase (Nb-CC4-ALP) fusion protein with good stability. A dual-emission system based ratiometric fluoroimmunoassay (RFIA) for quick and highly sensitive determination of 1-NAP was developed. Silicon nanoparticles (SiNPs) was used as an internal reference and for aggregation-induced emission enhancement (AIEE) of gold nanoclusters (AuNCs), while AuNCs could be quenched by MnO2 via oxidation. In the presence of ALP, ascorbic acid phosphate (AAP) can be transformed into ascorbic acid (AA), the later can etch MnO2 to recover the fluorescence of the AuNCs. Based on optimal conditions, the proposed assay showed 220-fold sensitivity improvement in comparison with conventional monoclonal antibody-based ELISA. The recovery test of urine samples and the validation by standard HPLC-FLD demonstrated the proposed assay was an ideal tool for screening 1-NAP and provided technical support for the monitoring of carbaryl exposure.