Efficient precious-metal utilization in electrocatalysis requires electrically conductive support architectures that enable electrocatalytic activity beyond conventional surface-limited designs. Herein, we demonstrate that solvent-assisted crystallization converts poly(3,4-ethylenedioxythiophene):poly(styrenesulfonate) (PEDOT:PSS) into a water-stable, highly conductive nanofibrillar matrix featuring nanoscale porosity and controlled swelling. During the subsequent electrodeposition of Pt nanoparticles (NPs), the highly porous nanofibrillar PEDOT:PSS moderately swells in aqueous media, allowing Pt ions to deeply infiltrate the polymer network and form uniformly dispersed Pt NPs throughout the entire film volume rather than on the surface alone. This volumetric nanoconfinement effect yields a markedly enlarged electrochemically active surface area (20 m2 gPt -1), rapid reactant permeability, and structural robustness under operating conditions. The resulting PEDOT:PSS-Pt nanocomposite exhibits greatly enhanced catalytic activity for hydrogen evolution and methanol oxidation reactions, outperforming conventional planar Pt architectures. This work establishes highly porous nanofibrillar PEDOT:PSS as a previously underutilized volumetric electrocatalyst scaffold and provides a general design strategy for maximizing precious-metal efficiency in electrocatalysis and water-splitting systems.
Diabetes mellitus represents one of the most prevalent chronic diseases worldwide, posing serious challenges to global health and healthcare sustainability. Conventional therapeutic strategies often face limitations such as poor bioavailability, frequent dosing, and lack of real-time glucose regulation. The emergence of smart nanotechnology offers transformative possibilities for the prevention, diagnosis, and management of diabetes within the broader framework of smart health and precision medicine. This review highlights recent advances in the design and application of nanomaterials for diabetes management, focusing on two key areas: drug delivery and glucose monitoring. Smart nanocarriers comprising polymeric, lipid-based, and metallic nanoparticles enable controlled and stimuli-responsive insulin release, improved pharmacokinetic profiles, and enhanced patient compliance. Concurrently, nano-enabled biosensors and wearable devices have revolutionized continuous glucose monitoring through superior sensitivity, selectivity, and integration with digital health platforms. The convergence of nanotechnology with artificial intelligence (AI), Internet of Medical Things (IoMT), and real-world health data further accelerates personalized diabetes care by enabling predictive monitoring and adaptive insulin therapy. Despite remarkable progress, challenges remain regarding clinical translation, long-term biosafety, and regulatory standardization. This review discusses these aspects comprehensively and provides future perspectives on integrating smart nanotechnology into sustainable, patient-centered diabetes management systems.
The unchecked and uncontrolled global spread of multidrug-resistant (MDR) bacteria is a serious challenge to healthcare and modern medicine, and demands diagnostic approaches that are rapid, sensitive, multiplexed, and reliable in point-of-care (POC) settings. At the interface of nanomaterials and aptamer-based biosensing, significant advances have been reported. The convergence of portable electrochemical sensing technologies, smartphone-based readout systems, and artificial intelligence (AI)- and machine learning (ML)-based data analysis is also playing a significant role in advancing this area. This perspective reflects on the most recent breakthroughs and translational developments in electrochemical nano-aptasensors for MDR bacterial detection, covering diagnostic targets and translation trends, nanomaterials advancements, aptamer engineering-integration, POC strategies and microfluidics platforms, and novel multimodal strategies that enhance accuracy, reliability, and efficiency in POC testing. Moreover, limitations and knowledge gaps in this area, as well as key considerations for sustainable development, large-scale manufacturing, and deployment of integrated electrochemical nano-aptasensors, are also highlighted. Electrochemical nano-aptasensors can pave the way for the transformation of MDR bacterial diagnosis, but need coordinated translational efforts for POC diagnostic advancements towards real-world applications.
Nanopore is a single-molecule technology for sensing biomolecules. Biomolecular interactions are essential biological processes that govern biological functions and therapeutic responses. However, high-resolution nanopore sensing of biomolecular interactions, such as protein-ligand interactions, remains challenging. In this study, we demonstrate that a YaxAB nanopore with LiCl-modulated electrostatic potential enables detection of molecular interactions of the BRD4 protein with histone peptides, as well as diverse small-molecule drugs, at the single-molecule level. Our electrical recordings and molecular dynamics simulations confirm that the oscillating dynamics of BRD4 within the funneled YaxAB nanopore generate two-level current transitions between narrow- and wide-pore regions. Using the parameters derived from dual-level dynamics and their signal decomposition, a YaxAB nanopore sensing approach enables the sensitive discrimination of BRD4-small-molecule drug complexes with a subtle mass difference as small as 2.5 Da. This near-atomic, high-resolution sensing capability of YaxAB nanopores may enable applications in single-molecule-based drug discovery, proteomics, and diagnostics.
Peripheral artery disease (PAD), characterized by progressive occlusion of peripheral arteries, is a major global health concern associated with high risks of ischemic complications and limb dysfunction. Endovascular stenting remains a primary therapeutic approach; however, the development of biodegradable vascular stents that offer both sufficient mechanical resilience and antithrombotic, anti-restenotic surfaces remains challenging, especially in highly deformable peripheral vessels. Herein, a 3D-printed biodegradable drug-eluting stent (DES) based on biofunctional silica-polycaprolactone nanocomposites and Janus surface nanoengineering is presented. Sol-gel-derived silica incorporation and extrusion-based 3D printing yield stents with tuned radial strength, elliptical struts that reduce flow disturbance, and enhanced support for endothelial regeneration. Janus nanoengineering is achieved through tantalum (Ta) plasma immersion ion implantation. The resultant nano-Ta-enriched luminal surface promotes human umbilical vein endothelial cell adhesion and proliferation. Meanwhile, the abluminal layer, comprising sirolimus/poly-L-lactic acid and nano-Ta, suppresses vascular smooth muscle cell proliferation, reduces platelet thrombosis, and minimizes the initial burst release of therapeutic agents. Comprehensive in vitro hemocompatibility and cytocompatibility studies, combined with in vivo evaluation in a PAD model, demonstrate improved patency, reduced neointimal hyperplasia, and favorable tissue responses. This 3D-printed, Janus-engineered DES represents a promising theragenerative platform for vascular tissue engineering.
Nanocellulose has long been studied as a bioactive material for tissue engineering; however, the mechanisms underlying its surface chemistry-mediated immune reprogramming remain unclear. Herein, we report a comprehensive multi-omics study of pristine cellulose nanocrystals (CNCs) and amide-functionalized CNCs (a-CNCs) to elucidate their 'nano-immune' interaction and impact on tissue-resident macrophages in vivo. Using integrated scRNA-Seq, bulk RNA-Seq, pharmacological inhibition, and histological profiling, we reveal that a-CNCs exhibit outstanding biocompatibility, showing no pro-inflammatory activation of macrophages across major organs within 14-day subacute window. In particular, a-CNCs exposure correlates with enhanced voltage-gated ion channel (KCa3.1 and Scn1b) and Stat6 signaling, while suppressing Nfkb-driven pro-inflammatory signals. This suggest that ion channel activation is strongly associated with M2 macrophage polarization. Moreover, a 28-day splenocytes profiling revealed no observable increase in CD4+/CD8+ T cells, suggesting non-adaptive immune response after a-CNC exposure. Concurrently, pseudotime mapping further discloses that a-CNC exposure preserves natural macrophage developmental trajectories across organ niches, while pristine CNCs induce mild M1-skewing in the spleen. In vitro validation confirms that a-CNCs intrinsically drive a pro-healing phenotype in macrophages, underscoring that macro-scale immune behavior can be transcriptionally triggered through nano-level surface chemistry of CNCs.
In this study, we developed UV-laser-induced carbon nanosphere/graphene (UV-LICNG) composites using a single-step ablation technique. This method employs UV-laser-induced forward transfer (UV-LIFT) to directly fabricate line-patterned UV-LICNG composites on silane-terminated polyurethane (S-PU) substrates with excellent mechanical properties. The unique structure of UV-LICNG, comprising conjugated carbon nanospheres and graphene with a large surface area, enables outstanding strain and humidity sensing performance. Owing to a separation-based sensing mechanism, the UV-LICNG-based strain sensor exhibits highly sensitive strain detection in the low-strain regime, achieving a high gauge factor (GF ≈ 146.5 within the 0-2% strain range), along with excellent linearity (R² ≈ 0.9906), rapid response and recovery times (29 ms and 31 ms, respectively), and exceptional durability over 3,000 stretching cycles at 0.2% strain. These attributes enable precise detection of subtle human motions and vocalization-induced strain signals. In addition, the intrinsic nano-micro porous graphitic structure of UV-LICNG imparts excellent humidity-sensing performance, characterized by fast response and recovery times (4.2 and 4.8 s, respectively), thereby facilitating reliable respiration monitoring and non-contact skin humidity sensing. The combined strain and humidity sensing capabilities, together with the simplicity and scalability of the UV-LIFT process, highlight the strong potential of UV-LICNG-based wearable electronics for continuous human health monitoring and multifunctional wearable sensing applications. The online version contains supplementary material available at 10.1007/s42114-026-01732-8.
Atomic force microscopy (AFM) enables label-free nanoscale imaging and nanomechanical profiling but remains constrained by low throughput, operator dependence, and variability in data interpretation. Artificial intelligence (AI) transforms AFM into a scalable and adaptive platform. Initially applied in materials science for super-resolution imaging, tip deconvolution, segmentation, and force-curve analysis, AI approaches are now being extended to biological AFM. These methods support robust denoising of soft matter maps, automated recognition of heterogeneous structures, and three-dimensional reconstruction of biomolecular assemblies. This review provides an end-to-end workflow of AI-enabled AFM─from probe optimization and adaptive control to multimodal data integration─highlighting advances relevant to mechanobiology and biomedical engineering. By surveying studies with amyloid fibrils, extracellular vesicles, membranes, and living cells, we show how AI-AFM convergence enhances reproducibility, throughput, and clinical utility. AI-driven AFM is poised to enable disease modeling, therapeutic screening, and precision diagnostics, establishing itself as a next-generation tool for biomedical discovery.
Plasmonic dimers are versatile platforms for manipulating light-matter interactions at the nanoscale, supporting hybridized modes such as capacitive plasmons (CPs) and charge transfer plasmons (CTPs), which are highly sensitive to the nature of the interparticle junction. However, these junctions have largely been restricted to noble metals, limiting fundamental understanding and design flexibility. Here, we report gold nanosphere dimers interconnected by a metal-semiconductor hybrid junction that enables selective regulation of plasmonic modes. Single-particle scattering measurements show that the hybrid junction, comprising metallic Ag pathways embedded within a high-permittivity AgI matrix, produces enhanced CPs and suppressed CTPs. Supported by electromagnetic simulations, we reveal that interfacial field localization driven by induced dipoles in AgI governs the mode selectivity by trapping oscillating surface plasmons and impeding long-range electronic conduction. This hybrid junction offers a tunable plasmonic platform, expanding opportunities in surface-enhanced Raman spectroscopy, optothermal therapeutics, nanophotonics, and optoelectronics that benefit from enhanced CP modes.
Titanium dioxide (TiO2) has evolved from a conventional photocatalyst into a sophisticated nano-platform that bridges environmental sustainability and biomedicine. This paper proposes a unified interfacial redox design framework that links the electronic-structure engineering of the TiO2 with the spatial control of its reactive oxygen species (ROS). In the environmental sector, we highlight advances in photocatalytic detoxification, such as the cleavage of organophosphates via Ag-modified TiO2, driven by doping and metal-support interactions. In the biomedical domain, TiO2 is framed as an active bio-interface capable of coordinative protein binding. We specifically examine the "moonlighting" protein dihydrolipoamide dehydrogenase (DLDH) as a model for stable, oriented biofunctionalization. By integrating RGD-targeting motifs, these hybrid systems enable integrin-directed, localized photodynamic effects. We further address critical toxicological considerations, emphasizing that TiO2 behavior is context-dependent and governed by particle size, crystallinity, and surface state. By synthesizing insights from catalysis and redox biology, this manuscript outlines principles for the rational design of safer, application-specific TiO2 technologies. This convergence supports a transition from non-selective oxidation toward predictable, spatially confined redox outcomes in both complex environmental matrices and physiological systems. This review outlines key mechanistic insights and proposes design principles for controlled and context-dependent TiO2 activity.
Achieving concurrent detection of high-priority polycyclic aromatic hydrocarbons (PAHs) in particulate matter (PM) using a single sensing platform remains challenging due to their low abundance, structural similarity, and coexistence in complex environmental matrices. Herein, we present a first voltammetric sensing platform, enabling simultaneous determination of benzo[a]pyrene (BaP), pyrene (Pyr), and fluorene (Fluo) using a single-chip screen-printed carbon electrode (SPCE) modified with polydopamine (PDA)-functionalized multiwalled carbon nanotubes (MWCNTs) decorated with CeO2 nanospheres. Structural and compositional features of the composite were investigated using field-emission scanning electron microscopy, transmission electron microscopy, X-ray diffraction, Raman spectroscopy, and X-ray photoelectron spectroscopy. The MWCNT/PDA/CeO2/SPCE exhibited excellent electrocatalytic activity toward the direct oxidation of BaP, Pyr, and Fluo in a methanol/water electrolyte containing LiClO4. This enhanced performance arises from the synergistic effects, including hydrophobic interactions, π-π stacking, efficient PAH adsorption, and improved electron-transfer kinetics. The sensor enabled simultaneous determination of BaP, Pyr, and Fluo with wide linear ranges (0.5-12, 0.1-10, and 7-60 μM, respectively), high sensitivities (6.99, 9.17, and 1.24 μA/μM, respectively), and low detection limits (0.22, 0.08, and 5.55 μM, respectively), along with excellent reproducibility (RSD < 4%) and selectivity against structurally similar PAHs. The practical applicability of the sensor was demonstrated by applying it to PM samples collected from on-road and different urban environments, which were fortified with known concentrations of the target analytes, yielding recoveries of 91.4-107.5% and confirming reliable performance in complex airborne matrices. Overall, the MWCNT/PDA/CeO2/SPCE platform enables a robust, sensitive, and selective strategy for simultaneous monitoring of multiple PAHs in environmental samples.
Airway obstruction resulting from both malignant and non-malignant etiologies is a growing challenge in pulmonary diseases and critical care medicine, particularly after the COVID-19 pandemic. Conventional silicone and metallic airway stents may be indicated in airway obstructions that lead to palliative relief, but they may lead to complications such as migration, inflammatory reaction to the adjacent tissue, and granulation tissue overgrowth. We conducted this animal pilot study to investigate the biocompatibility of a next-generation nanocomposite silicone airway stent, engineered with 3wt% hydrophobic nano-silica reinforcement. Innovative characteristics of the stent include improved biocompatibility and reduced mucus adhesion due to its hydrophobic properties. A refined stenting technique was applied to implant the stent in the trachea of two sheep models by assembling two endotracheal tubes, Ambu, and the stent. After a two-month follow-up, high-resolution computed tomography imaging, 3D virtual bronchoscopy, bronchoscopy, and biopsy of the tracheal wall were done. Histopathologic assessment demonstrated an inflammatory infiltrate dominated by lymphocytes, without stromal reactions, mucosal and submucosal thickening, or granulation, confirming a favorable tissue tolerance. These preliminary outcomes emphasize the stent's potential as a transformative therapeutic option; however, the study's limited sample size and absence of comparative controls highlight the necessity for further preclinical trials with quantitative airflow parameters to elucidate the clinical translatability of this innovative biomaterial solution for airway obstructions. Additionally, the findings of this study can address the unmet needs in managing complex airway obstructions, particularly for patients refractory to current therapeutic options in the future.
Developing an efficient and safe therapy necessitates a mechanistic understanding of the complex underlying pathology and manipulation of the multiple pathways at the molecular and genetic level. Network-based simulation of chronic myeloid leukemia (CML), a relatively well-understood cancer model, revealed the dynamics of simultaneously expressing pro-apoptotic BIM (Bcl-2 interacting mediator of cell death) and silencing pro-survival MCL-1 (myeloid cell leukemia-1) in combination with the breakpoint cluster region (BCR)-Abelson (ABL)-targeted tyrosine kinase inhibitor dasatinib. Viral/nonviral chimeric nanoparticles (ChNPs) composed of a BIM-expressing adeno-associated virus (AAV) core and a degradable polymeric shell that encapsulates MCL-1 siRNA (BIM/MCL-1 ChNPs) selectively killed BCR-ABL + CML cells in combination with dasatinib. In a mouse CML model, the BIM/MCL-1 ChNPs and dasatinib combination therapy suppressed proliferation of BCR-ABL + hematopoietic cells and prevented leukemic infiltration of organs. The enhanced anti-leukemic effect was further pronounced in an acute phase model of the disease. This study investigated a strategy of developing a versatile and tunable multimodal therapy assisted by a computational toolset that analyzes the molecular foundation of a disease and predicts therapeutic response. The interdisciplinary approach developed and validated in this study can be used in discovering new therapies for cancer and other diseases. [Image: see text] The online version contains supplementary material available at 10.1186/s40580-026-00543-3.
Radiation therapy induces DNA damage primarily through reactive oxygen species, leading to cancer cell apoptosis. However, intratumoral heterogeneity and spatial dose variations often result in the survival of polyploid giant cancer cells (PGCCs), a therapy-resistant subpopulation characterized by multinucleation, genetic instability, and stem-like features. Particularly in malignant breast cancer, PGCCs contribute to recurrence by adopting a dormant yet invasive phenotype. Despite their clinical relevance, reliable tools to identify or characterize these cells remain lacking. Here, we present a nanomechanical single-cell profiling platform that enables high-resolution mechanomics of radiation-induced PGCCs. Through integrated cytoskeletal imaging and nanoscale stiffness mapping, we identify a distinct mechanical dormancy state, marked by cortical actin remodeling, nuclear enlargement, and biomechanical stiffening. This dormant mechanotype is coupled with suppressed proliferation yet sustained expression of invasion-associated markers, representing a latent therapeutic threat. Our findings position mechanical dormancy as a mechanobiological hallmark of radiation resistance and propose a predictive framework for optimizing radiotherapy thresholds. This platform enables mechanotype-guided stratification and precision-targeted intervention in radiation-refractory cancer.
Alkali rare-earth fluorides have defined the golden era of upconversion nanoparticles (UCNPs) in biophotonics. Yet, their exceptional structural and chemical stability has raised concerns over long-term bioretention and potential chronic toxicity. Here, we present a strategy that bridges the gap between high-performance photonics and biological safety by simply substituting rare-earth ions with Zr4+ on the highly active [001] facet of LiREF4 (RE = Yb3+), which induces the formation of a biodegradable domain while preserving the structural integrity and luminescent efficiency of the native lattice. Density functional theory calculations reveal that this domain can stably host Zr4+ in the form of hydrolytically active zirconium fluoride complexes with coordination numbers ranging from six to eight. The configurational diversity of these complexes contributes to a significantly extended degradation lifetime, with four days in the bare form and over one month when coated with silica via the Stöber process. Our strategy establishes a new design rule for imparting biodegradability to highly efficient upconversion materials with day-scale degradation, thereby paving the way for clinically translatable optical imaging and therapeutic applications.
Ginkgo biloba L., a 'living fossil' with an evolutionary history spanning over 200 million years, occupies a prominent place in traditional medicine systems. Historical records, including the Chinese pharmacopoeia Ben Cao Gang Mu, document its use for treating various ailments related to the respiratory, circulatory, and cognitive systems. Modern pharmacological research has validated these traditional applications, identifying Ginkgo biloba extract (GBE) as a valuable phytomedicine for neurodegenerative, cardiovascular, and metabolic diseases. This comprehensive review aims to synthesize recent advances in GBE research, with a primary focus on literature from the past three years in GBE research, critically evaluating four key areas: 1) innovations in green extraction and purification technologies; 2) novel organelle-level mechanisms of action; 3) the development and application of nano-delivery systems for targeted therapy; and 4) clinical evidence and toxicological studies supporting GBE's therapeutic use and safety profile. This holistic review was conducted through a systematic literature search (covering 2003.8.15-2026.2.22) in databases including PubMed, Web of Science, and Scopus, using keywords related to GBE's extraction technologies, nano-delivery systems, organelle-level mechanisms, and clinical applications. The retrieved literature was screened and critically evaluated to synthesize recent advances, with a focus on identifying emerging trends, convergent evidence, and knowledge gaps across the four key areas outlined in the review's aim. Significant improvements in the sustainability and efficiency of GBE extraction have been achieved through recent technological innovations. (Deep Eutectic Solvent) DESs have emerged as tunable platforms for the selective and efficient extraction of bioactive compounds, while physical field-assisted methods enhance yields. Mechanistic studies reveal that GBE orchestrates cellular homeostasis by synchronously modulating mitochondrial bioenergetics, endoplasmic reticulum stress, and autophagic-lysosomal activity. Nano-delivery systems, such as liposomal nanoparticles and chitosan-coated carriers, effectively enhance the bioavailability and targeting precision of GBE's bioactive components, including quercetin. Clinically, standardized formulations like Ginkgo Diterpene Lactone Meglumine Injection (GDLI) show promise in improving cognitive outcomes in ischemic stroke and Alzheimer's disease, and in managing cardiovascular diseases. However, potential toxic side effects, primarily from ginkgolic acids (GA) and 4'-O-methylpyridoxine (MPN), necessitate rigorous quality control, with advanced methods like enzymatic degradation being developed for detoxification. GBE represents a promising, multifaceted therapeutic agent that effectively bridges traditional medicine and modern scientific innovation. Its broad-spectrum efficacy stems from the synergistic interactions of its bioactive constituents and their system-level regulatory mechanisms. To fully harness GBE's potential as a precision therapeutic agent, future research should focus on a coordinated technological strategy: employing green extraction methods such as DES to ensure sustainable and efficient compound isolation; developing standardized nano-formulations to overcome key pharmacokinetic limitations like low bioavailability and poor targeting; and applying multi-omics approaches to elucidate its organelle-level mechanisms of action. Validating these integrated innovations in large-scale clinical trials will be essential. Ultimately, the convergence of green chemistry, nanomedicine, and systems biology is key to translating GBE's multifaceted bioactivity into targeted and reliable clinical therapies.
Chimeric antigen receptor (CAR)-T therapy has led to remarkable advancements in the treatment of hematologic malignancies, encouraging extensive studies on its application to solid tumors and other diseases. However, the production of CAR-T cells is mostly achieved through viral transduction, which results in permanent CAR expression in T cells, potentially leading to unintended adverse effects. Here, we present a lipid nanoparticle (LNP) platform for mRNA delivery to human primary T cells, inspired by the human immunodeficiency virus (HIV) which naturally infects T cells. We perform multiple rounds of screening to sequentially optimize the structure and ratio of ionizable lipid in the base formulation, the ratios of HIV lipid components, and the type and ratio of PEG-lipid for CD3 antibody conjugation. Our HIV envelope-Inspired T cell transfection-Enhancing (HITE) LNP enables efficient generation of CAR-T cells with potent cytotoxic activity against cancer cells in vitro, demonstrating its potential for efficient CAR-T cell production.
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Montmorillonite (MMT) is a natural clay mineral and the major component of bentonite, and it possesses an exceptional swelling capacity, a high potential for adsorption, and excellent biocompatibility. Therefore, MMT has been identified as an attractive candidate for pharmaceutical applications in drug nano delivery systems. In recent years, research on MMT has also gained momentum because of its ability to improve the solubility, rate of dissolution, and bioavailability of poor-water-soluble drugs in polymeric nanocomposites. Although much research has been done on the formulation advantages of MMT, critical analysis of their translational progress and industrial uptake remains inadequate. This reviews an in-depth study of the role of MMT as a pharmaceutical excipient in nano drug delivery through a synergy of published work and patent landscape analysis. Patents on MMT-based formulations are sequenced and scrutinized to identify significant players in the industry, therapeutic applications, and emerging patterns of technology. In the comparative assessment of research publications vis-a-vis patents in the decade under review, evolving innovation patterns have of late drawn attention to scalable, multifunctional, clinically relevant MMT nanocomposites. Overall, the review indicates how MMT is gaining importance in pharmaceutical development and what it will provide for the future concerning rational design, regulatory convergence, and commercialization of MMT-embedded polymeric nano drug delivery systems.
DNA methylation at 5-methylcytosine (5mC) is crucial for embryonic development and cellular function, while aberrant patterns strongly drive disease onset and progression. Its reversible nature offers substantial therapeutic potential, emphasizing the need for precise, context-specific genome wide 5mC mapping. Conventional techniques such as bisulfite sequencing and ensemble biosensor assays are hindered by DNA degradation, amplification bias, high cost, and inability to resolve single-molecule structural and mechanical effects of methylation. This review examines advances in single-molecule biophysical methods (nanopore sensing, smFRET, optical/magnetic tweezers, and AFM) that provide direct, label-free/minimally invasive 5mC detection, along with quantitative insights into DNA conformation, mechanics, and protein-DNA interactions. These techniques complement traditional methylome mapping by linking genomic localization to molecular mechanisms. Emerging machine-learning approaches are revolutionizing analysis, particularly in nanopore sensing, while promising applications in smFRET, tweezers, and AFM address throughput and reproducibility challenges. Their convergence promises scalable, high-resolution epigenetic profiling, advancing precision epigenomics toward clinical application.