Compared with oxidoreductase mimics, hydrolase-like nanozymes can circumvent the interference from redox species in real matrices, exhibiting unique superiority for biochemical sensing, but insufficient catalytic efficiency and lack of specific response to targets when no bioreceptor (aptamer, antibody, etc.) is employed greatly restrict their advanced analytical applications. To solve this predicament, here we employed a ligand engineering strategy to modulate the phosphatase-mimetic activity of Nd(III)-coordinated phenylboronic acid (PBA) derivatives and achieved anti-interference and sensitive detection of quinolone antibiotics in dairy products based on matched interaction and ligand-optimized catalytic efficiency. By tailoring the ortho-substituted PBA ligand, precise activity control was obtained through adjusting surface Lewis acidity and substrate adsorption jointly. Quinolones could specifically suppress the activity of Nd-PBA via strong hydrogen bonding, π-π stacking, and coordination interactions, allowing the nanozyme to function as both recognition unit and signal transducer without the need for external bioreceptors. According to the principle, high-performance sensing of ciprofloxacin as a quinolone model was validated. By leveraging the synergy of specific interaction and hydrolase-mimicking catalytic nature, the established approach exhibited good anti-interference capability in complex matrices. The ligand-modulated catalytic activity provided sufficient sensitivity for detecting the analyte in dairy products. Our attribution not only develops a new hydrolase mimic with engineerable phosphatase-like catalytic characteristics via ligand programming but also offers a rarely explored approach for selectively detecting quinolone antibiotic residues, inspiring future design of bioreceptor-free nanozyme sensing systems for broader applications.

Lactylation is a widespread protein modification with important regulatory roles in health and disease. Emerging evidence also links metabolite lactylation to disease onset, progression, and treatment response. It is estimated that an uncharacterized pool of lactoyl-metabolites exists in vivo, according to its formation mechanism. To mine this pool, we combined knowledge-driven prediction with stable isotopic labeling-based high-resolution mass spectrometry (SIL-HRMS). A predictive library of 151 lactoyl-metabolites was generated by expanding 20 basic amino acids (AAs) to their upstream and downstream metabolites. To improve detection, we developed a pair of new hydroxyl-reactive labeling reagents, 3,5-(dimethylamino)-2,4,6-triazine benzene-1-chlorine (Tmt-aycl-Cl) and its deuterium form d12-Tmt-aycl-Cl. Using HRMS, 95 lactoyl-metabolites were putatively identified across 14 tissue types, 64 structures of which are reported for the first time. Representative lactoyl-metabolites were chemically synthesized and characterized to verify some of the annotations. We then developed a pseudotargeted metabolomics workflow for semiquantitative profiling in a diabetic kidney disease (DKD) mouse model. DKD mice showed elevated lactate in serum and the kidney and widespread dysregulations of lactoyl-metabolites. Many lactoyl-metabolites correlated strongly with the urinary albumin-to-creatinine ratio, suggesting biomarker potential for DKD. This integrated strategy expands the catalog of lactoyl-metabolites and provides a platform for investigating their biological significance.

Antibody aggregation is a widespread phenomenon that frequently occurs when antibodies interact with solid surfaces. It can significantly compromise antibody functionality, e.g., reduced antigen-binding affinity, altered pharmacokinetics, and increased immunogenicity, making it a major concern in therapeutic and diagnostic applications. Therefore, understanding the molecular interactions between antibody structures and material surfaces is critical for optimizing antibody drug formulation, storage, and filtration processes. In this study, we investigated the surface-induced aggregation behavior of two antibodies─IgG-1 and IgG-4─using polypropylene (PP) and poly(vinyl chloride) (PVC) films as representative hydrophobic and hydrophilic polymer surfaces, respectively, with a combined experimental and computational approach. In situ sum frequency generation (SFG) vibrational spectroscopy was used to monitor interfacial protein structures at varying concentrations, while solvent-accessible surface area (SASA) calculations provided structural and hydropathological insights into antibody-surface interactions. Significantly stronger SFG signals were observed for IgG-1 than for IgG-4 on both PP and PVC surfaces, across both low and high (1.2 and 12 mg/mL) concentration conditions. IgG-1 showed marked orientational rearrangement upon surface contact, in contrast to the relatively stable configuration of IgG-4. SASA calculations further supported these observations by revealing a greater exposure of hydrophobic regions in IgG-1, which likely contributes to its enhanced surface affinity and fouling propensity. Together, these results provide molecular-level insights into antibody-surface interactions and highlight molecular determinants that govern protein aggregation on polymeric filtration materials. This work offers guidance for the rational design of low-fouling biomaterials and improved antibody handling strategies in biomanufacturing and therapeutic applications.

Diabetes is a highly prevalent chronic disease worldwide, and its early diagnosis is crucial for long-term monitoring and effective management. Sensitive identification and detection of glycated hemoglobin A1c (HbA1c), a key biomarker that reliably reflects average blood glucose levels over the past 2-3 months, play an important role in the long-term monitoring of diabetes. Here, we present a fluorescent and aptamer-based highly sensitive biosensor method for HbA1c detection employing a newly designed, target-triggered, and movable toehold-assisted proximal catalytic hairpin assembly (mt-pCHA) amplification strategy. The recognition of HbA1c by the aptamer triggers the release of DNA strands to activate the mt-pCHA process, in which the assembly hairpins are arranged in close proximity on the preassembled Y-shaped scaffolds to enhance their local concentrations for accelerating reaction kinetics. Furthermore, the introduction of a movable toehold optimizes the strand displacement pathway and suppresses the background leakage while preserving the catalytic efficiency. With the HbA1c-triggered initiation of mt-pCHA, numerous quenched fluorescent hairpins are unfolded in the assembly process, leading to significantly magnified fluorescence recovery for ultrasensitive detection of HbA1c. Experimental validation demonstrates a strong linear response over the range of 5 ng/mL-50 μg/mL for HbA1c with a detection limit of 4.66 ng/mL. The assay exhibits negligible cross-reactivity toward hemoglobin (Hb) and performs reliably in diluted human whole blood hemolysate samples. These results highlight its potential as a versatile biosensing platform for the sensitive detection of low-abundance protein biomarkers.

Chiral discrimination of amino acid enantiomers is critical in food safety and biomedical fields due to their distinct biological activities and potential toxicity. This study presented a simple strategy using the achiral metal-organic framework MIL-101(Cr)-NH2 as a direct fluorescent sensor for the visual enantioselective recognition and quantification of lysine and arginine. The probe operated through a "turn-on" response driven by differential host-guest interactions without requiring any chiral modification. It exhibited exceptional enantioselectivity, with a fluorescence intensity ratio (FL/FD) of 107 for lysine and a reverse selectivity (FD/FL = 1.66) for arginine, achieving nanomolar detection limits. Mechanism studies revealed that the enantiomeric preference originated from distinct hydrogen bonding and electrostatic interactions within the MOF pores. The sensor demonstrated practical utility in accurately detecting l-Lys in commercial yogurt samples with satisfactory recoveries (91.7-103.7%). Furthermore, by integrating Rhodamine B, a smartphone-based colorimetric platform was developed for the on-site visual assessment of enantiomeric excess (ee) values, showing a linear correlation (R2 = 0.9943) between the RGB ratio and the ee value of Lys. This work highlights that readily available achiral MOFs can serve as cost-effective and versatile platforms for chiral discrimination, offering a promising alternative to conventional chiral sensing systems in food analysis and biomedical diagnostics.

Herein, by formamide self-condensation, an innovative Fe-Cu dual-atom nanozyme (FeCu-N-C) is presented, engineered with adjacent metal sites within a nitrogen-doped carbon scaffold, that exhibits synergistically enhanced peroxidase-, oxidase-, and laccase-like activities. This multienzymatic performance far exceeds that of single-atom Fe-N-C and Cu-N-C controls. Density functional theory (DFT) calculations reveal that the proximity of Cu sites induces an upshift in the Fe d-band center from -1.52 eV to -0.88 eV, optimizing substrate adsorption and providing a mechanistic rationale for the observed catalytic synergy. Leveraging this multienzymatic platform, we developed a single-material colorimetric sensor array capable of discriminating five distinct pesticides in complex matrices. These pesticides differentially modulate the triple-mimetic activities through compound-specific interaction mechanisms, as validated by DFT analysis. Integrated with an artificial neural network (ANN), the system achieved 100% identification accuracy. This work establishes a pioneering paradigm for the rational design of multienzyme mimics and underscores their potential for high-precision environmental monitoring.

Influenza A (H1N1), which is a highly contagious respiratory pathogen, poses a serious public health threat. Its effective control necessitates rapid and accurate detection methods. Here, we developed a novel quantum dot (QDs) nanobeacon-based biosensor that synergistically integrates the precise target recognition and trans-cleavage activity of CRISPR/Cas13a with the isothermal, enzyme-free signal amplification power of catalytic hairpin assembly (CHA) for sensitive detection of H1N1 RNA. This system achieves dual signal amplification, enabling detection with a remarkably low limit of detection (LOD) of 95.2 aM within 40 min. Excellent specificity distinguishes H1N1 from other common influenza viruses. Crucially, the programmability of crRNA endows the method with broad versatility. Beyond influenza virus detection, it was successfully applied to SARS-CoV-2 RNA detection, achieving an LOD of 87.0 aM, demonstrating its potential for diverse pathogen diagnostics. These results collectively demonstrate the high performance and adaptability of this diagnostic platform, highlighting its significant promise for future applications in the rapid and sensitive detection of diverse viral pathogens.

Despite notable advances in molecular diagnostics, developing ultrasensitive biosensors for biomarker quantification remains a critical unmet need. Herein, we address this gap by engineering lanthanide metal-organic frameworks (Ln-MOFs) as artificial receptor aptamers (ARAs) via rational ligand design and guided optimization. Our lead Eu-TPTC Ln-MOF ([1,1':4',1″]-terphenyl-3,3″,5,5″-tetracarboxylic acid), optimized for maximal surface negativity, binds the cationic biomarker (cardiac troponin I, cTnI) with highly efficient electrostatic affinity. This strong affinity couples synergistically with specific target recognition to enable robust, binding-triggered signal transduction. A phosphate-mediated cascaded amplification strategy achieves a 7250-fold fluorescence enhancement and a cTnI limit of detection (LOD) of 1 pg mL-1. This performance meets clinical requirements for early diagnosis and significantly outperforms conventional ELISA in sensitivity (12.3-fold), speed, and cost-effectiveness. We establish three core ARA design principles: (i) a rigorous experimental structure-property framework linking ligand functional groups to MOF surface zeta potentials; (ii) multiscale interface engineering combining molecular docking and fluorescence assays; and (iii) an affinity-driven signal-leverage strategy. This label-free platform enables miniaturized, field-deployable diagnostics, critical for resource-limited settings. Notably, it provides a general framework for MOF-based ARAs, extends to diverse biomarkers, and lays a solid foundation for the development of highly responsive, enzyme-free biosensors for ultrasensitive biomarker quantification.

Extracellular vesicles (EVs) have emerged as a promising class of liquid biopsy biomarkers, and programmed death-ligand 1-positive EVs (PD-L1+ EVs) have been proven to accurately predict patient response to tumor immunotherapy. Therefore, rapid and sensitive detection of PD-L1+ EVs is of great importance for improving treatment efficiency and promoting clinical application. Here, we developed a microchip-based gauge (MicroGauge) for rapid (<30 min) and highly sensitive detection (up to 4.8 × 104 particles/mL) of the purified salivary PD-L1+ EVs by reading a visible distance signal, thereby facilitating real-time prediction of immunotherapy efficacy. In this method, the purified salivary PD-L1+ EVs are efficiently captured via a one-step immune reaction and then injected into a MicroGauge with a dual pressure-division structure and an inclined capture structure. MicroGauge guides the immune complex to arrange in a monolayer close-packed array, finally forming a visual distance signal and achieving a good sensitivity. Using this MicroGauge, it was found that tumor patients had a higher level of salivary PD-L1+ EVs than healthy individuals. Moreover, to evaluate the clinical application potential, an immunotherapy clinical trial was conducted in oral cancer patients. By comparing the changes in the signal length before and after patients received immunotherapy, we found that the signal length of those with a good treatment response continued to shorten during the treatment process, while nonresponders showed a trend of first decreasing and then increasing. This result indicates that MicroGauge can effectively predict the efficacy of immunotherapy by rapidly and highly sensitively detecting the level of PD-L1+ EVs.

Comprehensive xenometabolome characterization is essential for understanding the effects of xenobiotics in biological systems. This study presents a multidimensional analytical workflow integrating orthogonal chromatographic separations, trapped ion mobility spectrometry (TIMS), high-resolution mass spectrometry and biotransformation-informed data processing to address xenometabolome assessment challenges. Zebrafish larvae exposed to 4-Methylbenzotriazole (4-MeBT) were used as a challenging case study. TIMS dimension provided orthogonal experimental evidence for isomer annotation, with inverse reduced mobility (1/K0) supporting conjugation site assignment for the dominant O-S-4MeBT and O-G-4MeBT isomers. The combination of TIMS with the Parallel Accumulation Serial Fragmentation (PASEF) acquisition further reduced spectral complexity, enhanced signal-to-noise ratio, and improved MS/MS coverage (70%), generating high-quality analytical evidence crucial for structural elucidation. To leverage these analytical dimensions, we developed a data processing strategy that leverages in-silico-based suspect screening and biotransformation-informed nontarget screening. In this regard, we introduce two novel frameworks; the "Building Blocks" (BB) concept which interprets unknown bio-TPs as modular assemblies of parent- and pathway-derived substructures, and the "Spectral Characteristics Knowledgebase" (SCKB), which use known biotransformation MS/MS motifs to provide structural insights and facilitate unknown identification. Our results demonstrated the identification of all previously known 4-MeBT bio-TPs with enhanced confidence (O-Sulfate- and O-Glucuronide-4MeBT) and the discovery of 29 new bio-TP features across 12 bio-TP classes, highlighting its efficacy in unraveling complex xenobiotic metabolism. Among these, a putative dimerization product (4-MeBT-263) was reported for the first time in zebrafish. Overall, this workflow has the potential to advance the understanding of bio-TP formation and detoxification processes in xenometabolome studies.