Real-time monitoring of pork freshness using polyvinyl alcohol/modified agar multilayer gas-sensitive labels.

Accurate consumer perception of food packages should provide real-time feedback on any changes inside food packaging. Hence, a new multilayer gas-sensitive label (POA-12) was prepared using a layer-by-layer pouring method for simple, visual, and real-time detection of pork's freshness, while the front side was developed by immobilizing red carbon dots and fluorescein isothiocyanate in POA as indicator for volatile nitrogen, and the back side was created using bromothymol blue in POA as pH indicator. The swelling index of the multilayer gas-sensitive labels reduced from 159.19% to 148.36%, and the tensile strength increased from 25.52 MPa to 42.61 MPa. In addition, the POA-12 multilayer label showed a red-to-yellow fluorescence change as TVB-N increased from 6.84 to 31.4 and a yellow-brown-to-blue-green color change as pH increased from 5.74 to 7.24 when detecting pork samples. Thus, it provides dual-indicator monitoring that improves the accuracy and reliability of assessing the freshness of high-protein products.

Enhanced Photothermal Activity of Nanoconjugated System via Covalent Organic Frameworks as the Springboard.

The development of nanomaterials with high photothermal conversion efficiency has been a hot issue. In this work, a novel photothermal nanomaterial is synthesized using Prussian blue nanocubes (PBNCs) as the photothermal active substance and covalent organic framework (COF) as the substrate. The as-prepared COF@PBNCs show a high photothermal conversion efficiency of 59.1%, significantly higher than that of pure PBNCs (32.5%). A new circuit path is generated with the combination of COF, which prevents the direct combination of thermal electrons and holes, as well as enhances the nonradiation transition of PBNCs. Besides, the imine groups on COF as the coordination and reduction agent allow the in situ growth of PBNCs, and the dense micropores of COF as the ideal heat conduction channels can also be the potential factors for the enhanced photothermal property. The photothermal property of COF@PBNCs is further used in the construction of immunosensor for the detection of furosemide (FUR). With the help of handheld thermal imager, the concentration of FUR can be easily read, thus shedding a new light in the construction of visual sensor for simple and low-cost point-of-care testing.

DNA Nanomachine-Driven Heterogeneous Quadratic Amplification for Sensitive and Programmable miRNA Profiling.

Inspired by the molecular crowding effect in biological systems, a novel heterogeneous quadratic amplification molecular circuit (HEQAC) was developed for sensitive bimodal miRNA profiling (HEQAC-BMP) by combining an MNAzyme-based DNA nanomachine with an entropy-driven catalytic hairpin assembly (E-CHA) autocatalytic circuit. Utilizing ferromagnetic nanomaterials as the substrate for DNA nanomachines, a biomimetic heterogeneous interface was established; thus, a localized molecular crowding system was created that can elevate the local reaction concentration and accelerate the molecular recognition process for a significant threshold signal. Simultaneously, the threshold signal undergoes further amplification by E-CHA and is transformed into a chemical signal, enabling a colorimetric-fluorescence bimodal signal readout. The HEQAC-BMP enables miRNA detection from 10 aM to 10 nM with detection limits of 3.7 aM (colorimetry) and 4.8 aM (fluorometry), respectively. Moreover, the design principle and strategy of HEQAC-BMP can be customized to address other critical viruses or diseases with life-threatening and socioeconomic impacts, enhancing healthcare outcomes for individuals.

Boosting ultralong chemiluminescence for the self-powered time-resolved immunosensor.

The construction of an immunosensor based on ultralong chemiluminescence is challenged due to the shortage of highly efficient initiator for long and stable catalysis. Herein, the heterogeneous Au/Pt@CuO/Cu2O catalyst was used to investigate the structure-activity relationship, while Au/Pt significantly promotes the activity of CuO/Cu2O to catalyze H2O2 and thus produces ·OH and O2•- radicals in highly alkaline solutions, resulting in the strong and long chemiluminescence in the reaction with luminol (10 mL, more than 4 min with 1 µg catalyst). By using the Au/Pt@CuO/Cu2O as the label in the immunoassay, the strong and long chemiluminescence could initiate the photocurrent of the photoelectrochemical (PEC) substrate, and the luminescence time could influence the photocurrent extinction time, thus a self-powered time-resolved PEC immunosensor was developed to detect furosemide, showing a linear relationship between the extinction time and the logarithm of concentrations from 10-3 to 1 µg/L. This work not only experimentally verifies that the Pt-O-Cu bond in heterogeneous catalysts breaks the pH limitation of the Fenton reaction, but also realizes the chemiluminescence for self-powered time-resolved immunosensor, thereby expanding the portable applicability of chemiluminescence in food safety inspection, health monitoring, and biomedical detection without external light source.

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.

Advances in immunoassay-based strategies for mycotoxin detection in food: From single-mode immunosensors to dual-mode immunosensors.

Mycotoxin contamination in foods and other goods has become a broad issue owing to serious toxicity, tremendous threat to public safety, and terrible loss of resources. Herein, it is necessary to develop simple, sensitive, inexpensive, and rapid platforms for the detection of mycotoxins. Currently, the limitation of instrumental and chemical methods cannot be massively applied in practice. Immunoassays are considered one of the best candidates for toxin detection due to their simplicity, rapidness, and cost-effectiveness. Especially, the field of dual-mode immunosensors and corresponding assays is rapidly developing as an advanced and intersected technology. So, this review summarized the types and detection principles of single-mode immunosensors including optical and electrical immunosensors in recent years, then focused on developing dual-mode immunosensors including integrated immunosensors and combined immunosensors to detect mycotoxins, as well as the combination of dual-mode immunosensors with a portable device for point-of-care test. The remaining challenges were discussed with the aim of stimulating future development of dual-mode immunosensors to accelerate the transformation of scientific laboratory technologies into easy-to-operate and rapid detection platforms.

PEI-coated Prussian blue nanocubes as pH-Switchable nanozyme: Broad-pH-responsive immunoassay for illegal additive.

Nanozymes are commonly used in the construction of immunosensors, yet they are generally susceptible to pH condition, which greatly hindered their practical use. To break the limitation of pH conditions, polyethyleneimine-coated Prussian blue nanocubes (PBNCs@PEI) were synthesized as the pH-switchable nanozyme, which can show peroxidase-like and catalase-like activity in acidic and alkaline condition, respectively. Besides, the modification of PEI can largely improve the catalytic activity of PBNCs. Herein, the pH-switchable catalytic property of PBNCs@PEI was used to construct the dual-mode immunosensor for the detection of illegal additive, rosiglitazone. In acidic condition, PBNCs@PEI showed excellent peroxidase-like activity, which can trigger the colorimetric reaction of Au nanostars with TMB2+/CTAB. In alkaline condition, the catalase-like activity of PBNCs@PEI prevailed, thus the decomposition of H2O2 can generate O2 to initiate the aerobic oxidation of 4-chloro-1-naphthol (4-CN), which can decrease the fluorescence intensity of 4-CN. Based on the competitive immunoassay, both the localized surface plasmon resonance wavelength shift of Au nanostars and the fluorescence intensity change of 4-CN were quantitatively related with rosiglitazone concentration, thus shedding a new light on the construction of broad-pH-responsive immunosensor. Besides, a smart device was developed to transfer the chroma value of Au nanostars into the RSG concentration, making this sensor a promising method in on-site and point-of-care detection.

Handheld Platform for Sensitive Rosiglitazone Detection: Immunosensor Based on a Time-Based Readout Device.

The detection of rosiglitazone (RSG) in food is of great importance since the excessive intake of RSG could cause adverse effects on the human body. Although liquid chromatography-mass spectrometry and gas chromatography-mass spectrometry are the preliminary methods for the detection of hazardous materials in food, they are not suitable for point-of-care or on-site detection. Herein, a time-based readout (TBR) device with an application software (APP) controlled by a smart phone was developed for the sensitive and selective immunoassay of RSG. The homemade TBR device was based on a two-electrode system, where the immune molecule-modified glassy carbon electrode was used as the bioanode, and Prussian blue-modified FTO was used as the cathode. By using Au-modified octahedral Cu2O with high catalytic activity as mimetic peroxidase, an insulating layer was generated on the cathode by catalyzing 4-chloro-1-naphthol (4-CN) into benzo-4-chlorohexadienone (B4Q). The time to reach a fixed potential varied indirectly with the concentrations of RSG and was recognized by the APP, while the electrochromic property on the cathode was also correspondingly changed. Under optimum conditions, both the square root of the time and the chroma value of the electrochromism exhibited linear responses for the detection of RSG ranging from 5 × 10-10 to 5 × 10-7 g/L, while the limits of detection were 8.2 × 10-11 and 1.3 × 10-10 g/L, respectively. With easy operation and portability, this TBR device showed a promising application for point-of-care monitoring of hazardous materials in food or the environment.

"Dual-Signal-On" Integrated-Type Biosensor for Portable Detection of miRNA: Cas12a-Induced Photoelectrochemistry and Fluorescence Strategy.

The abnormal expression of microRNA (miRNA) can affect the RNA transcription and protein translation, leading to tumor progression and metastasis. Currently, the accurate detection of aberrant expression of miRNA, particularly using a portable detection system, remains a great challenge. Herein, a novel dual-mode biosensor with high sensitivity and robustness for miR-21 detection was developed based on the cis-cleavage and trans-cleavage activities of Cas12a. miRNA can be combined with hairpin DNA-horseradish peroxidase anchored on a CdS/g-C3N4/B-TiO2 photoelectrode, thus the nonenzymatic amplification was triggered to form numerous HRP-modified double-stranded DNA (HRP-dsDNA). Then, HRP-dsDNA can be specifically recognized and efficiently cis-cleaved by Cas12a nucleases to detach HRP from the substrate, while the remaining HRP on HRP-dsDNA can catalyze 4-chloro-1-naphthol (4-CN) to form biocatalytic precipitation (BCP) on the surface of the photoelectrode, and thus the photocurrent can be changed. Meanwhile, the trans-cleavage ability of Cas12a was activated, and nonspecifically degrade the FQ-reporter and a significant fluorescence signal can be generated. Such two different kinds of signals with independent transmission paths can mutually support to improve the performance of the detection platform. Besides, a portable device was constructed for the point-of-care (POC) detection of miR-21. Moreover, the dual-mode detection platform can be easily expanded for the specific detection of other types of biomarkers by changing the sequence of hairpin DNA, thereby promoting the establishment of POC detection for early cancer diagnosis.

Toll-like receptor 4 promotes high glucose-induced catabolic and inflammatory responses in chondrocytes in an NF-κB-dependent manner.

Diabetes is an independent risk factor for knee osteoarthritis (OA), and hyperglycaemia-induced inflammation is considered to play an important role in their connection. The Toll-like receptor 4 (TLR4) regulates inflammatory responses in several pathological conditions including diabetes and OA. However, its role in diabetes-associated OA is poorly understood. In this study, we found that TLR4 expression was higher in OA cartilage from patients with type 2 diabetes mellitus (T2DM) than that from non-T2DM patients. Similarly, its expression was induced in primary mouse chondrocytes treated with high glucose, which suggests that TLR4 upregulation in T2DM-associated OA cartilage may originate from hyperglycaemia stimulation. We further discovered that TLR4 promoted high glucose-induced catabolic and inflammatory responses in chondrocytes, and mechanistically, these effects could be explained by the exacerbated activation of the transcription factor nuclear factor kappa B (NF-κB) pathway, since its inhibition by Bay 11-7082 abrogated TLR4 effects on high glucose-treated chondrocytes. Taken together, these findings may reveal a promotive role of TLR4 in regulating hyperglycaemia-induced catabolism and inflammation in T2DM-associated OA, and also implicate that TLR4 inhibition might be of therapeutic significance in treating T2DM-associated OA.