Understanding biomineralization processes requires experimental strategies that can correlate morphological, chemical, and structural information across scales. However, bridging microscale morphological and chemical descriptions with nanoscale molecular information remains a major experimental challenge. The multimodal and multiscale workflow developed in this work integrates scanning electron microscopy with energy-dispersive X-ray spectroscopy (SEM/EDX), infrared microspectroscopy (μFT-IR), and atomic force microscopy-infrared spectroscopy (AFM-IR) performed sequentially on the same specimen. It addresses the main challenges associated with multimodal analysis, including sample preparation compatibility, measurement chronology, and region-of-interest colocalization through a straightforward colocalization and image registration procedure. When applied to a breast biopsy sample containing microcalcifications, this experimental strategy enabled the resolution of both inter- and intra-deposit chemical heterogeneities. By integrating complementary spectroscopy datasets, this workflow provides a chemical description of mineralized tissues spanning the micro- to the nanoscale, thereby linking the specimen's global and local physicochemical properties. Beyond this application, this study provides a methodological framework for investigating mineralization pathways and hierarchical organization in complex biomineralized systems.
Pathogenic bacteria are related to clinical infections and high morbidity, and they pose significant diagnostic challenges for rapid testing. Loop-mediated isothermal amplification (LAMP) is a fast, convenient, and highly sensitive nucleic acid assay but remains challenging in complex matrices as well as in strategies for fast and accurate result reading. In this study, a cationic platinum nanocluster-based loop-mediated isothermal amplification platform with pretreatment by magnetic nanoparticles was innovatively developed for the assay of methicillin-resistant Staphylococcus aureus (MRSA) in clinical samples. Concanavalin A-modified magnetic nanoparticles (ConA-MNPs) showed excellent separation performance and stability, with a capture efficiency (CE) >92% in HEPES. Meanwhile, we also innovatively combined nanozymes of cationic CS-PtNCs with LAMP to explore a new form of LAMP signaling output mode, enabling on-site, rapid readout of assay results. Furthermore, the excellent peroxidase-like activity of CS-PtNCs enables colorimetric signal output of LAMP results for accurate and quantitative detection of MRSA. The platform enables dual-mode detection (visual and colorimetric) of MRSA in clinical CSF samples in less than 1 hour, with a limit of detection (LOD) as low as 20 CFU per mL. In the future, this method is expected to enable field applications in clinical pathogen diagnosis and disease prevention, and could evolve into a universal detection platform by simply replacing components such as primers.
We report a tetrahedral DNA platform that positions a catalytic hairpin assembly within an optimal spatial confinement window, thereby maximizing target-triggered amplification relative to background leakage. AS1411-mediated targeting and a β-actin reference channel further enable selective, internally calibrated imaging and sensing of intracellular microRNA-21.
Ensuring food safety requires straightforward methods for assessing food quality to protect public health. The decomposition of meat/fish proteins produces ammonia and biogenic amines (BAs), which can be sensitively detected and are essential for evaluating food safety during storage, transport, and consumption. We have designed and synthesized three ESIPT-exhibiting luminescent probes [3,3″-bis(benzo[d]thiazol-2-yl)-[1,1':4',1″-terphenyl]-4,4″-diol (B1), 3-(benzo[d]thiazol-2-yl)-[1,1'-biphenyl]-4-ol (B2), and 3,5-bis(benzo[d]thiazol-2-yl)-[1,1'-biphenyl]-4-ol (B3)] derived from 2-(2'-hydroxyphenyl)benzothiazole (HBT), simply by varying the number and position of benzothiazolate units on the hydroxy-phenyl backbone. The probability of intramolecular proton-transfer of the phenolic O-H group may be increased in the presence of two benzothiazolate units occupying both ortho positions of the central O-H (designated as B3) due to the possibility of increased intramolecular hydrogen bonding. Such disposition of the functional groups facilitates excited-state intramolecular proton transfer (ESIPT) and thus results in a higher pKa. In contrast, the presence of only one benzothiazolate unit at the ortho position of the O-H (B1 and B2) relatively imparts a lesser extent of intramolecular hydrogen bonding, leading to a lower pKa value. The lower pKa values of B1 and B2 make them particularly suitable for pH sensing, enabling multiple fluorescence color changes at different pH levels. Moreover, these probes have been tested for amine sensing, achieving limits of detection (LOD) of 1-4.5 µM and 0.5-1.5 µM for ammonia and hydrazine, respectively, with ratiometric response. The strip-based detection of fish spoilage has been demonstrated for real field applications.
In this work, we developed a novel approach driven by molecular crowding conditions combined with padlock probe amplification (MCC-PPA) to detect the rpoB 531 (TCG> TTG) mutation in Mycobacterium tuberculosis...
Peroxynitrite (ONOO-) is a highly reactive nitrogen species that plays pivotal roles in diverse physiological activities and pathological events. However, the sensitive and organelle-specific detection of ONOO- in complex biological environments remains a considerable challenge. In this study, we developed BMZ-ON, a novel near-infrared (NIR) "turn-on" fluorescent probe for the selective detection of mitochondrial ONOO-. Upon reaction with ONOO-, BMZ-ON undergoes a specific cleavage process to release BMZ-OH, accompanied by a marked enhancement of NIR fluorescence. The probe exhibited high sensitivity, with a detection limit of 95 nM, together with excellent selectivity toward ONOO- over other biologically relevant reactive species. In addition, co-localization studies demonstrated its effective mitochondrial targeting capability and good membrane permeability. BMZ-ON was further applied to visualize endogenous ONOO- fluctuations in living cells and zebrafish embryos. These results indicate that BMZ-ON provides an effective platform for monitoring mitochondrial ONOO- and may facilitate further studies on the involvement of ONOO- in mitochondrial dysfunction and related diseases.
Developing a low-cost, highly sensitive, and accurate fluorescence-based approach for Cu2+ detection is crucial for reducing heavy metal pollution in industrial effluent. Here, we report a newly synthesized Schiff base europium metal-organic framework (Eu-MOF) and demonstrate its use for turn-off fluorescent detection of Cu2+. A Schiff base ligand derived from 2-aminoterephthalic acid and 2-hydroxynicotinaldehyde was coordinated with Eu3+ to construct the framework, enabling a simple and rapid sample pretreatment procedure. The material exhibited pH-dependent fluorescence behavior. With the concentration range of 2.5 to 500 mg L-1, a linear response to Cu2+ was observed. The limit of quantification (LOQ) was 2.5 mg L-1, while the limit of detection (LOD) was 0.83 mg L-1. The method was effectively performed on vinegar, tea, fruit juice, and water samples, yielding spike recoveries of 84.5%-100.9% and relative standard deviations (RSDs) of 1.7%-4.8%. This method can be used to determine Cu2+ in various matrices.
The discrimination of structurally similar biothiols remains a critical challenge in clinical diagnostics, as conventional nanozyme-based sensor arrays often suffer from limited signal diversity and poor discriminative ability. To address this, a colorimetric sensor array using two rationally designed chiral carbon dots (CDs) was constructed. The role of chirality was primarily reflected in regulating the catalytic heterogeneity and enriching the multi-channel response diversity rather than direct enantioselective recognition. The two chiral CDs possess markedly distinct oxidase-like and laccase-like activities. This disparity in catalytic activity effectively doubles the number of response channels, thereby enriching the cross-reactive signal patterns. The CDs catalyzed the oxidation of three chromogenic substrates, namely 3,3',5,5'-tetramethylbenzidine (TMB), 2,2'-azinobis(3-ethylbenzothiazoline-6-sulfonic acid) (ABTS), and the 4-aminoantipyrine/2,4-dichlorophenol (4-AP/2,4-DP) coupling system in the absence of H2O2, producing distinct colorimetric responses. The addition of thiols elicits a differential inhibition of this activity in a degree-dependent manner, thereby inducing characteristic changes in the colorimetric readout. The developed array enabled accurate simultaneous identification and quantitative analysis of four thiols, including glutathione (GSH), L-cysteine (Cys), homocysteine (Hcy), and thioglycolic acid (TGA), by extracting RGB values using a smartphone color picker app and combining them with principal component analysis (PCA), hierarchical cluster analysis (HCA), and supervised machine learning. The k-nearest neighbor (KNN), support vector machine (SVM), and random forest (RF) classifiers achieved overall classification accuracies ranging from 95.83% to 97.22%, which further confirmed the robustness and reliability of the proposed sensing strategy. PCA score plots showed well-separated clusters for all analytes over the concentration range of 1-100 μM, while HCA further confirmed 100% classification accuracy. Furthermore, the first principal component (PC1) exhibited good linear correlation with GSH and TGA concentrations in a wide linear range of 1-50 μM, and the detection limit for all thiols was 1 μM. Successful application in serum and urine samples highlights its potential for smartphone-based clinical disease diagnosis.
Aflatoxin contamination, especially aflatoxin B1 (AFB1), is a major global health concern. There is an urgent need to develop rapid, highly sensitive and specific sensing platforms. In this study, two fluorescence aptasensors were developed for AFB1 detection. The first used quantum dots (QDs) combined with aptamers as molecular recognition elements, along with RecJf exonuclease amplification and Fe3O4 nanoparticles for magnetic separation. In the absence of AFB1, the aptamer stably hybridizes with the cDNA-QDs conjugate, thereby shielding its 5' end from cleavage by RecJf exonuclease. Upon AFB1 binding, the duplex dissociates, exposing the aptamer's 5'-single-stranded region to RecJf-mediated hydrolysis-resulting in AFB1 release and enabling catalytic recycling and cascade signal amplification. It achieved a detection range of 0.01-100 ng mL-1 and a limit of 2.2 pg mL-1 (3σ/s). To further reduce interference and improve sensitivity, a second sensing system based on upconversion nanomaterials (UCNPs) was developed. This system, also using RecJf exonuclease amplification, achieved a broader range of 0.001-100 ng mL-1 and a lower detection limit of 0.56 pg mL-1. The QDs-based system required 90 min for detection, whereas the UCNPs-based system achieved detection in just 10 min. The two detection methods, particularly the UCNPs based one, enable specific identification of AFB1. Both aptasensors were successfully applied to the determination of AFB1 in spiked wheat flour and corn flour samples, with recoveries ranging from 89% to 111.6%, demonstrating their applicability to AFB1 analysis in cereal products. By replacing the aptamer, the proposed aptasensor can be further extended for the detection of other biological or chemical targets.
At present, the detection methods of tumor markers rely on ELISA, gene sequencing, and other technologies. However, these methods require specialized equipment and technical personnel, and the low concentrations of early tumor markers limit their sensitivity. This study developed a method for detecting colorectal cancer tumour markers. It uses a test strip and isothermal amplification to detect low concentrations of tumour markers. The test strip is easy to use and cost-effective. The test strip developed in this study can detect isothermal amplification products in 3 min, and the detection limit for PPIA is 5 ng mL-1. This method is expected to play a significant role in early cancer detection, providing an efficient and convenient approach for tumor marker detection.
Acoustic tweezers use acoustic radiation force and acoustic streaming generated by ultrasound fields to achieve contactless and relatively low-damage manipulation of biological targets, including cells, bacteria, extracellular vesicles, and other micro-/nanoscale objects. In biosensing workflows, they mainly function as modules for sample preparation, target enrichment, sorting, reaction organization, and mass-transfer enhancement before downstream signal readout. This review summarizes the fundamental mechanisms, representative device configurations, and integration strategies of acoustic tweezers, with emphasis on how their analytical roles change with target scale. Radiation-force-based focusing and sorting are most effective for cells and bacteria, whereas nanoscale targets increasingly require localized streaming, interfacial acoustic fields, microbubble oscillation, or vortex-based manipulation; molecular assays mainly benefit from enhanced mixing, mass transfer, and droplet organization. Recent applications are evaluated according to the analytical bottleneck addressed, quantitative performance improvement, sample conditions, and remaining limitations. Acoustic tweezers are further compared with optical tweezers, dielectrophoresis, magnetic manipulation, inertial microfluidics, and deterministic lateral displacement using common workflow-level criteria. They offer a balanced combination of label-free operation, biocompatibility, throughput, and microfluidic integration, but are not universally superior in precision or nanoscale control. Clinical translation remains limited by force-streaming competition, complex-matrix field distortion, acoustic heating, device-to-device variation, cross-talk with downstream readout, and insufficient validation in minimally processed clinical samples. Future efforts should focus on scale-specific multiphysics design, closed-loop workflow integration, standardized manufacturing and reporting, and clinically defined validation to advance acoustic tweezers from laboratory prototypes toward deployable point-of-care testing systems.
Understanding the behaviour of lipids during digestion of food and formulations can enable more rational design of lipid-based materials to enhance the oral delivery of drugs and nutrients. Traditionally, light microscopy and electron microscopy have been used to 'watch' digestion at the droplet level, while small angle X-ray scattering has more recently given structural insights at the ensemble level, however none of these techniques provide information on local changes in chemical composition of a digesting fat droplet. In this study, we used synchrotron Fourier transform infrared microspectroscopy (s-FTIRM) as a chemical imaging technique to map the distributions of milk components and their digestion products in individual fat globules to obtain a better understanding of the interfacial characteristics and chemical specificity behind the structural changes that occur during digestion. The results showed localisation of fatty acids at the core and the oil/water interface of digesting fat globules and the presence of fatty acid-calcium soaps, which exist both as isolated aggregates and are partially accumulated at the interface. This proof-of-concept study highlights the novel application of s-FTIRM as a label-free, chemically-specific imaging tool to resolve the spatial distribution of digestion products, such as free fatty acids and fatty acid-calcium soaps, within and around individual milk fat globules. This approach provides new insights into the localised chemical changes during lipid digestion at the single droplet level.
This work presents the development, characterization, and partial analytical method validation of a non-enzymatic electrochemical sensor based on a nanocomposite comprising iron oxide magnetic nanoparticles (MNPs), phosphorylated cellulose nanofibers (pCNF),...
Selenium and cadmium, as trace elements widely present in the environment, their stress mechanisms in vivo, involve multiple aspects, such as oxidative stress, metabolic disruption, and apoptosis, among other aspects....
Sensitive and accurate monitoring of arginase activity is significant for clinical diagnosis. Herein, a dual-enhanced aggregation-induced electrochemiluminescence (ECL) biosensor was constructed based on triple-ligand-protected gold nanoclusters (AuNCs), sequentially coating 6-aza-2-thiothymine-capped AuNCs with L-arginine (Arg) and tetraoctylammonium bromide (TOAB). This ligand-engineering strategy restricted molecular vibrations and rotations while enhancing electron transfer efficiency, leading to a stepwise increase in ECL efficiency from 23% to 30% and finally to 71%. The hydrolysis of Arg, catalyzed by arginase, leads to dramatic quenching in the dual-enhanced ECL signal, forming the basis for quantitative detection of arginase activity. It achieved an ultra-low detection limit of 1.23 × 10-7 U mL-1 with a broad linear range from 1 × 10-6 to 0.02 U mL-1. The system showed excellent selectivity, stability, and reproducibility and was successfully applied to determine arginase activity in human serum. Triple-ligand AuNCs with dual-enhanced aggregation-induced ECL open a new avenue for constructing high-performance ECL detection platforms.
The advancement of microfluidic technologies has enabled more rapid analysis and reduced cost per reaction through system miniaturization, demonstrating great potential in diagnostics, partiThe advancement of microfluidic technologies has enabled...
Surface-enhanced Raman scattering (SERS) enables real-time analysis of volatile organic compounds (VOCs) and other gaseous molecules using portable Raman devices, offering faster measurements compared to GC-MS. To facilitate this, numerous solid SERS substrates have been developed and applied across multiple fields. However, they often require complex fabrication due to poor adsorption of gases onto solid surfaces leading to high production costs and inconsistencies in sensitivity and reproducibility. In contrast, colloidal nanoparticles are less expensive to produce and established facile synthesis methods help their broader implementation. In this study a simple approach has been conceptualised whereby gaseous analytes are bubbled directly into colloidal nanoparticles. This was achieved by pumping air into a flask containing the volatile analyte, which in turn was bubbled into a vial containing colloidal nanoparticles and measured with a portable Raman spectrometer. Initially, 3-mercapto-hexanol (3MH) was successfully measured using this method. 3MH was chosen due to the method originally being developed to measure thiols associated to underarm odour. Subsequently, 3MH was used to optimize the air flow rate by testing at eight different flow rates. The results demonstrated that flow rate influenced the timing of analyte spectral appearance as seen by the rate of change in peak height at 634 cm-1 for 3MH, with the most reproducible results observed at 50 mL min-1. The technique was further evaluated with two other analytes: 2-methyl-3-mercapto-pentanol (2M3MP) and indole. The latter needed the addition of an aggregating agent (NaCl) to the nanoparticles. These findings indicate that this approach offers a promising alternative for detecting gaseous analytes using colloidal nanoparticles.
The clustered regularly interspaced short palindromic repeats (CRISPR) and CRISPR-associated proteins (Cas) system enables sensitive and specific biomolecular detection due to its programmability, high fidelity, and signal amplification. Herein, a novel universal CRISPR/Cas12a-G4 DNAzyme-TMB (Cas-GT) enzymatic biosensing platform was constructed. This platform regulates the catalytic function of G4 DNAzyme through the trans-cleavage activity of Cas12a, achieving an "off/on" response of the TMB-H2O2 enzymatic signal for nucleic acid, protein, and metal ion targets. In proof-of-concept experiments, the detection limit of Cas-GT for in vitro transcribed SARS-CoV-2 RNA reached as low as 100 aM and it distinguished clinical positive from negative patients with good diagnostic performance (AUC = 0.9420). It is also suitable for protein targets, enabling quantitative analysis of prostate-specific antigen (PSA) within the range of 0-100 ng mL-1, with results highly consistent with clinical chemiluminescence immunoassay (CLIA). For small-molecule targets, Cas-GT exhibited good universality, achieving quantitative detection of Hg2+ within the range of 0.06-4 ng mL-1, with recovery rates of 93%-107.4% in spiked river water samples, showing no significant difference from ICP-MS (P = 0.9968). In summary, Cas-GT is a simple, sensitive, rapid, and label-free enzymatic biosensing platform with significant potential for clinical biomarker detection and environmental pollution monitoring.
Parathyroid hormone (PTH) is essential for calcium-phosphate homeostasis and bone metabolism. Abnormal PTH levels are closely associated with chronic kidney disease and several types of cancer. Current mainstream immunological PTH detection methods cannot achieve single-cell analysis, and dual-signal detection strategies are scarce. Herein, we designed a dual-signal nano-microelectrode for single-cell PTH detection by modifying gold-deposited nanopipettes with ferrocene (Fc)-labeled DNA-complementary DNA double strands and 6-mercaptohexanol (6-MCH). Under optimized conditions, square-wave voltammetry (SWV) showed a linear response to PTH in the range of 1-500 pg mL-1 (R2 = 0.99), and EIS exhibited linearity in the range of 1-1000 pg mL-1 (R2 = 0.98). The sensor demonstrated high selectivity against common intracellular interferents and good electrode-to-electrode consistency. Using three-dimensional micro-manipulation technology, minimally invasive detection of individual cells was achieved, and significant changes in electrochemical signals before and after cell penetration were revealed. Furthermore, the intracellular PTH concentration gradually decreased with prolonged extracellular Ca2+ incubation, consistent with the regulatory mechanism of calcium-sensing receptors. This dual-mode nano-micro electrode is a new type of tool that enables rapid, low-cost, label-free and highly specific single-cell PTH analysis.
