Nature Nanotechnology

The Wnt/{beta}-catenin pathway is constitutively active in most colorectal and other cancers. Tankyrase (TNKS) promotes Wnt signalling by PARylating AXIN, a rate-limiting scaffold subunit of the {beta}-catenin destruction complex, and its inhibition is a validated strategy for pathway downregulation. Existing small-molecule TNKS inhibitors that target the catalytic PARP domain suffer from off-target effects across the PARP family. Here, we present an alternative approach that targets the substrate binding domains of TNKS using a short TNKS-binding peptide (TBP) presented multivalently on DNA nanostructures. Such a strategy may be required to effectively disrupt the function of targets such as TNKS, which is known to form high-order assemblies in the cell. We engineered DNA nanostructures with two distinct geometries: a 2D triangle (~100 nm) displaying 27 copies of TBP, and a compact 3D tetrahedron (~10 nm) displaying 2 TBP copies. We show that both nanostructures assemble efficiently, can be functionalised with TBP in high yields, and retain binding to TNKS protein in vitro. DNA nanostructures are efficiently internalised to the cytoplasm by HeLa Kyoto and colorectal cancer cells, with TBP functionalisation enhancing rather than hindering uptake. In HeLa cells, both triangle-TBP and tetrahedron-TBP downregulated Wnt signalling to a similar extent despite an order-of-magnitude difference in TBP copy number, while the same concentration of free TBP had no effect. This finding likely reflects the two functions of the DNA nanostructures - intracellular delivery and multivalent display - whereby the smaller nanostructures more efficiently internalise the TBP ligand but have lower valency. These results establish DNA nanostructures as a modular, tuneable platform for multivalent inhibition of intracellular clustered targets such as TNKS.

Biological processes are often regulated through reactions involving supramolecular protein complexes organized with nanoscale precision, such as the apoptosome that activates procaspase-9 through defined stoichiometry to induce apoptosis. Here we report designer CASCAD (CARD Assembly on SCAffold DNA), a self-assembled DNA nanoplatform that spatially organizes caspase recruitment domains (CARDs), exhibiting catalytic intracellular caspase-9 activation. By functioning like a natural apoptosome, CASCAD enhances caspase-9 oligomerization and promotes valency-dependent enzymatic activation. Further, after incorporating targeting aptamers and membrane-interacting peptides for cellular uptake and cytosolic delivery, CASCAD has successfully reconstituted apoptosome function and downstream apoptotic signaling in in-vitro 2D and 3D cultures. Our molecular design paradigm establishes CASCAD as versatile and programmable DNA nanomaterial for reconstructing signaling pathways driven by diverse supramolecular complexes, including inflammasomes and myddosomes, providing a general chemical framework for engineering cellular signaling and directing cellular function and fate.

Rapid DNA transport limits the temporal resolution of solid-state nanopore sensing. Here we show that breaking pore symmetry through geometric inclination produces exceptionally prolonged DNA residence without surface functionalization or external trapping. For 500-base-pair double-stranded (ds) DNA ,13 nm diameter SiNx pores in tilted membranes exhibit a distinct long-lived population with a mean dwell time of 11.1 seconds at 600 mV bias, exceeding previously reported dsDNA dwell times by at-least 4 orders of magnitude. Controlled pore tilting reproduces the population of prolonged events, whose prevalence and duration depend on ionic strength. TEM-EELS based thickness measurements exclude substantial membrane-thickness differences, while continuum and atomistic simulations support a mechanism coupling electric-field redistribution to off-axis DNA motion, increased pore-wall contact and suppressed axial mobility. These findings establish geometric inclination as a passive design variable for engineering molecular residence in solid-state nanopores and provide a route towards longer observation windows for single-molecule sensing.

The nuclear pore complex (NPC) is a massive protein system that controls all nucleocytoplasmic transport via a dynamic network of intrinsically disordered proteins called FG-Nups. We developed an NPC-mimicking DNA origami nanostructure that self-assembles into a tetrameric, octagonal nanopore with an inner diameter that, through addition of DNA oligonucleotides, can be tuned in a user-defined fashion from 57 to 66 nm - mimicking the contracted and dilated states of NPCs. We demonstrated that these origami nanopores can dilate within minutes and recontract at slower timescales. Both contracted and dilated pores can be inserted into lipid bilayers, where they maintain their conformation and allow transmembrane transport of fluorescent proteins. With outer diameters up to 87 nm, these are, to our knowledge, the largest DNA origami nanopores inserted into lipid membranes. Functionalization of the pores with the FG-Nup Nsp1 reduces non-specific cargo diffusion across the lipid membrane while facilitating translocation of the transport receptor Kap95, demonstrating transport selectivity much like biological NPCs. The work establishes a size-tunable platform to dissect nuclear transport mechanisms, with potential applications for engineering programmable artificial channels.

Efficient intracellular delivery of neoantigen-encoded mRNAs is central to the development of effective cancer vaccines and immunotherapies. However, how the nanoscale geometry of delivery carriers impacts mRNA delivery remains poorly understood, largely because conventional delivery platforms do not readily permit systematic variation of carrier architecture while maintaining comparable chemical composition. Here, we develop a modular RNA-DNA hybrid nanoplatform that enables controlled investigation of geometry-dependent mRNA delivery. Guided by a unified design principle, we construct a series of two- and three-dimensional nanostructures with programmable architectures and systematically compare their delivery performance using consistent functional readouts. We further investigate how mRNA embedding strategies and aptamer-mediated functionalization influence intracellular delivery efficiency. By providing precise control over carrier geometry, this platform establishes a framework for elucidating structure-function relationships in mRNA delivery and offers a versatile strategy for engineering targeted nucleic acid delivery systems.

Antibody-DNA oligonucleotide conjugates (AOCs) are widely used for molecular assembly and cellular analysis, yet current approaches for generating these conjugates often rely on nonspecific chemistries that produce heterogeneous products. Here, we present two complementary strategies for generating site-specific AOCs using covalent DNA-linking HUH endonucleases. In one approach, recombinant antibodies are genetically fused to HUH-tags to enable direct, site-specific DNA conjugation. In the second, off-the-shelf antibodies are indirectly linked to HUH-tags using a photocrosslinkable Protein G-HUH fusion, enabling covalent Fc-directed attachment. Both strategies yield homogeneous AOCs while preserving antigen binding affinity. We apply these conjugates to a DNA-based mechanochemical assay, termed rupture-and-deliver tension gauge tethers (RAD-TGTs), which converts receptor-mediated adhesion forces into intracellular delivery of a fluorescent oligonucleotide payload. By tuning duplex stability, we define adhesion dynamics across multiple mechanical regimes. Using HER2- and beta1-integrin-targeting AOCs, we identify receptor-specific adhesion signatures and uncover cooperative interactions between receptor systems in a panel of cancer cell lines. Dual-color probes enable multiplexed single-cell mechanical phenotyping, and application to primary NK cells reveals dose-dependent responses to integrin modulators. These results establish a generalizable platform for site-defined AOC generation and for quantitative, high-throughput measurement of receptor-mediated adhesion dynamics.

Virus-like particles (VLPs) are non-infectious, multiprotein nanostructures that mimic the architecture of viruses while lacking genetic material. Their structural versatility and safety profile make them promising platforms for biomedical applications, including vaccine development, targeted drug delivery, and diagnostics. Among various functionalization strategies, click-chemistry has emerged as a precise and efficient method for attaching bioactive molecules to VLP surfaces. In this study, we report the production and functionalization of VLPs derived from the feline panleukopenia virus (FPV), a parvovirus closely related to the well-characterized canine parvovirus (CPV). We demonstrate successful expression, assembly, and purification of FPV VLPs, followed by their modification with fluorophores, peptides, oligonucleotides, and proteins using copper-free click-chemistry. Furthermore, using cryo-EM, we structurally characterize FPV VLP functionalized with antigenic peptides, providing detailed insights for further development of such vaccine prototype. Finally, we demonstrate that our FPV VLP specifically transduce feline and human cells expressing transferrin receptor type 1 (TfR), thus validating their potential as a unique platform for future nanotechnological and therapeutic applications.

Messenger RNA (mRNA) therapeutics have transformed vaccination and protein replacement strategies, yet efforts to improve their performance have focused largely on sequence engineering, nucleotide modification, and delivery vehicles. Here we show that mRNA function can be controlled by rational design of higher-order RNA architectures. We develop self-assembling mRNA origami (mRNA-OG), a class of unimolecular RNA nanostructures that encode protein-coding sequences within higher-order, programmable, and compact nucleic acid architectures. Using computational design and experimental validation, we demonstrate that mRNA-OG folds into well-defined nanostructures while remaining translationally competent in mammalian cells. Although folded mRNA-OG recruits ribosomes comparably to unfolded constructs, it produces lower protein output, indicating that RNA architecture can directly influence translational efficiency. The compact geometry of mRNA-OG also enhances encapsulation by cationic lipid delivery systems, suggesting a structural route to improved cargo packaging. Beyond its effects on translation, mRNA architecture modulates innate immune recognition. In primary human dendritic cells, folded and unfolded mRNA-OG elicit distinct cytokine programs and differential activation of stress-response pathways, including a modest induction of the integrated stress response that is not fully explained by canonical PKR signaling. Our results establish programmable structure as a new design parameter for mRNA therapeutics that can affect its functionality, delivery properties, and immune sensing.

Sophisticated biomolecular functions often arise from large, precisely organized architectures, motivating efforts to construct increasingly complex structures through programmed nucleic acid self-assembly. Although RNA offers a richer repertoire of structural motifs and biological functions than DNA, RNA nanostructures constructed to date have remained substantially less complex and less scalable than their DNA counterparts, most notably DNA origami and DNA bricks, which rely on large libraries of synthetic single-stranded (ss) DNA strands. Directly adapting these strategies to RNA, however, faces two major barriers: (i) the high cost of producing large libraries of distinct synthetic ssRNA strands, and (ii) the limited stability of ssRNA under conditions commonly used for assembling multicomponent nucleic acid nanostructures. Here we present the double-stranded RNA (dsRNA) bricks approach, which addresses both barriers by adopting principles inspired by natural RNA systems: (i) many distinct RNA components ("bricks") are encoded within a single precursor transcript and released by enzymatic processing, (ii) each brick's predominantly double-stranded nature enhances stability. The resulting dsRNA bricks self-assemble through programmable branched kissing-loop interactions. Using this approach, we constructed complex two- and three-dimensional RNA nanostructures with more than 100 distinct components, representing, to our knowledge, the largest and most compositionally complex RNA nanoarchitectures reported to date. Overall, the dsRNA bricks approach establishes a scalable and robust framework for constructing complex RNA nanostructures with compositional and architectural sophistication approaching that of DNA-based systems, thereby opening new opportunities for programmable RNA materials and RNA-based devices.

Molecular transport through a nanopore determines the information that can be recovered from a translocation signal, yet it remains difficult to control in conventional solid-state nanopores. Rapid translocation reduces the information content and fixed nanopore geometries limit the dimensionality of the signal. Here, we control molecular transport through the development of the pipette-elastomer interfacial nanopore (PEIN), a dynamically reconfigurable solid-state nanopore which addresses these limitations. A PEIN is formed by depressing a glass nanopipette into a soft elastomer, progressively constricting its aperture and enabling continuous control over aperture size, while retaining the simplicity and favourable noise characteristics of glass nanopipette sensing. Using the dynamic aperture size control, DNA velocities could be controlled over more than a twofold range, with dwell times two orders of magnitude greater than observed in glass nanopipettes. DNA-origami rulers further revealed a progressive reduction in polymer velocity during translocation, indicating that hydrodynamic drag alone is insufficient to model forces on the DNA polymer. Finally, by using single- and double-stranded DNA and gold nanoparticles as molecular standards, we demonstrate reversible, size-selective molecular gating with sub-nanometre control. These results establish the PEIN as an accessible platform for controlling molecular transport and probing the relationships among biopolymer conformation, nanoscale confinement and translocation dynamics.

Optically addressable spins, traditionally studied in semiconductors and, more recently, in (bio)chemical systems, are central to quantum technologies, yet existing platforms lack scalable and accessible site-specific programmability. Here, we show that DNA can serve as a functional nanoscale scaffold for optically addressable spin systems. By incorporating flavin chromophores into synthetic oligonucleotides, we generate spin-correlated radical pairs (SCRPs) that can be manipulated by radiofrequency (RF) fields and read out using optically detected magnetic resonance (ODMR). Pulsed ODMR resolves spin dynamics on sub-microsecond timescales under ambient conditions, while DNA sequence design enables atomically precise tuning of both the ODMR response and the associated spin chemistry with single-base resolution. DNA secondary structure provides an additional layer of functionality: duplex formation inverts the pulsed ODMR contrast, indicating a switch in the spin multiplicity of the SCRP precursor. The synthetic accessibility and chemical programmability of oligo-nucleotides as hosts for optically addressable spins are demonstrated through a series of proof-of-concept applications, including sensing, programmable SCRP positioning, and spin-enhanced molecular beacons. Our results establish DNA as a versatile scaffold for engineered spin systems, providing a platform for future applications ranging from quantum sensing and programmable spin arrays to bioimaging and RF-controlled molecular switches for gene regulation.

Homeostasis is a fundamental hallmark of living organisms, arising from the complex interplay between biochemical reactions and regulatory feedback systems. Reconstituting such self-regulating behaviour in minimal synthetic cells enables continuous, persistent operation of biochemical reactions for extended amount of time. In this work, we demonstrate a minimal homeostatic synthetic cell capable of autonomous flux regulation using DNA nanotechnology and bottom-up synthetic biology. Our homeostatic architecture consists of Giant Unilamellar Vesicles (GUVs) equipped with gated DNA nanopores, encapsulated in vitro transcription (IVT) machinery, and an RNA degradation system. We achieve homeostasis under varying external chemical stimuli specifically varying concentrations of rNTPs by implementing a negative feedback loop between rNTP influx and RNA production. In our system, DNA nanopores facilitate the influx of rNTPs from the external environment, driving internal transcription. Crucially, the transcription process generates RNA "blockers" designed to bind and gate the DNA nanopores, thereby attenuating further rNTP influx. Our system is dynamic as encapsulated RNases slowly degrade the RNA blockers, allowing the pores to reopen as blocker concentration goes down. We first characterise the functionality and gating efficiency of the DNA nanopores using both pre-synthesised and in situ produced DNA and RNA blockers. We then demonstrate that rNTP flux through these pores is sufficient to drive IVT within the GUVs. Finally, by integrating these modules, we demonstrate robust homeostasis: the system maintains a steady-state level of RNA production for up to 16 hours. By harnessing the controllability of negative feedback loop, we demonstrate thresholding of the homeostasis level using single-stranded regulator DNA. This work establishes a versatile framework for engineering adaptive and self-sustaining responsive nanomaterials and synthetic cell chassis.

Precise drug delivery within anatomically complex tissues demands systems capable of both active navigation and deep tissue access, properties that have remained difficult to reconcile in existing nanocarriers and microrobots. Here we introduce Dynabots, a dynamic microrobotic assembly constructed from multifunctional nanoparticles covalently linked by thermally cleavable molecular connectors. This nanoparticle-rich architecture enables the integration of magnetic, imaging, and therapeutic components while preserving a high content of functional material. Collective assembly imparts enhanced magnetic responsiveness and maneuverability, enabling controlled navigation through tortuous biological environments. Upon exposure to mild thermal stimuli, the assemblies undergo programmed disassembly, releasing individual nanoparticles that can diffuse through tissue for localized therapeutic action. We establish the programmable transitions, biocompatibility, and therapeutic efficacy of this process across in vitro and in vivo models, including real-time fluoroscopic guidance within anatomically realistic phantoms and live rodent and porcine systems. By integrating magnetic control, reconfigurable architecture, and stimulus-triggered disassembly, Dynabots unite navigational precision with tissue permeability, providing a versatile platform for adaptive and deep-tissue drug delivery.

Single-walled carbon nanotubes (SWCNTs) show promise for optical biosensing, imaging, and drug delivery, but turning them into safe, precision nanomedicine tools requires understanding how nanotube surface chemistry dictates recognition and processing by cells. Like other nanomaterials, carbon nanotubes acquire a biomolecular corona on contact with biological fluids, and corona identity is increasingly recognized as central to sensor performance and drug delivery efficacy. However, whether corona identity also governs the intracellular fate of carbon nanotubes remains largely unknown. Here, we show that the single-stranded DNA wrapping of (6,5)-enriched single-walled carbon nanotubes reprograms their protein corona, intracellular trafficking, and macrophage response. By profiling (AT)15, (GT)15, and (CT)15 wrapped SWCNTs, we show that the wrapping sequence programs both the protein corona and the resulting proteostatic stress on macrophages. Photoluminescence imaging and confocal Raman measurements reported that (AT)15 is internalized the most yet leaves the proteome and nanotube structure largely undisturbed, whereas (CT)15, taken up the least, undergoes the most aggressive intracellular degradation and drives the highest oxidative and proteostatic stress. Corona proteomics indicated that all three tested nanotubes form coronas with distinct functional identities that are responsible for divergent intracellular routes. Time-resolved intracellular proteomics combined with functional assays resolved how the host cell reorganizes its biomolecular complexity over time, including oxidative outputs, aside from a sequence-independent core response involving particle engagement, phagosomal sorting, and lysosomal processing. These findings provide mechanistic insight into nanomaterial-cell interactions and the wrapping sequence as a tunable, nucleotide-level design handle for controlling the intracellular fate of carbon nanomaterials, with potential implications for safe and effective nanomedicine platforms.

Nanomedicines that perform well in conventional cell cultures often fail to translate into patients, in part due to the inability to reproduce the protein corona, and hence the biological identity, that nanoparticles acquire within the tumor microenvironment (TME). Here, we establish patient-derived colonic organoids (PDCOs) as a platform for profiling nano-bio interactions under physiologically tumor-relevant conditions using graphene quantum dots (GQD) and Silicon Quantum Dots (SiQD) as material-distinct probes. By combining spectral flow cytometry, confocal imaging, dynamic light scattering, and multi-spectral dimension-reduction analysis of single-cell uptake, we resolve how nanoparticle identity, uptake, and intracellular fate diverge between conventional culture and patient tissue. Conventional two-dimensional culture overestimated internalization by two-fold relative to ex vivo PDCOs, while TME-specific proteins directed nanoparticles to distinct organoid subpopulations. Exploiting this, we engineer the protein corona with tumor-specific proteins to redirect SiQDs to chemoresistant cells, from 3.8% to 70%, an 18-fold increase in targeted delivery to the cells that drive therapy resistance. Internalization was also model-dependent. PDCOs and fibroblasts favored clathrin/dynamin (DNM1)-mediated endocytosis into lysosomes, whereas cancer cells favored caveolae (CAV1)-mediated entry that bypasses lysosomal degradation. This distinction determines whether a nanocarrier is degraded or delivered intact. Collectively, these findings turn the protein corona into a programmable design tool and establish PDCOs as a next-generation platform for precision nano-oncology.

Biomarker analysis requires detecting analyte classes that span nucleic acids, proteins, small molecules, and metabolites, yet testing remains fragmented across target-specific assays and instruments. Here we report a molecular information-transduction strategy that converts target recognition across molecular classes into a common, error-tolerant DNA code readable by nanopore sequencing. Target recognition triggers a hybridization chain reaction that generates concatemers containing periodically repeated 10-nucleotide target-specific barcodes. A matched-filter decoder exploits this periodicity and the linear scaling of read length with match count to reject spurious matches by two to three orders of magnitude. Multi-class detection is demonstrated for a small molecule (ATP), two cardiovascular-associated microRNAs and thrombin in singleplex and multiplexed assays. By separating molecular recognition from sequence readout, this architecture provides a modular framework for converting heterogeneous analytes into a shared, redundancy-encoded signal for high-fidelity molecular sensing.

Fluorescence microscopy is increasingly used to measure molecular organization at length scales where labeling, photophysics and sample preparation can dominate quantitative accuracy. Reference standards are therefore needed that combine defined nanoscale geometry with a protein-like environment and compatibility with biological imaging. Here we introduce circular tandem repeat protein as programmable protein standards for benchmarking sub-10-nm fluorescence microscopy. Using genetic code expansion and bioorthogonal labeling, we generated compact protein rings carrying up to six labeling sites. Photoswitching fingerprint analysis revealed geometry-dependent localization accumulation and blinking kinetics, demonstrating that short-range fluorophore interactions can be assessed as measurable benchmark parameters. DNA-PAINT confirmed accessible docking sites and programmed valency at the single-particle level. We further established recombinant tethering and genetically encoded membrane display, extending the standards to cellular environments. cTRP PicoRulers were also compatible with expansion microscopy. Together, cTRPs provide a modular protein-based platform for evaluating molecular-scale imaging performance in purified and cellular environments.

Selectively perturbing pathogenic membranes while preserving host-cell integrity is a fundamental challenge in the design of membrane-active biomaterials. This challenge is particularly acute for enveloped viruses because their lipid membranes are derived from host cells and therefore share many of the same molecular constituents. Here, we demonstrate that engineered nanochirality provides a structural parameter for distinguishing virus-organized membrane nanodomains from host-cell membranes. Histidine-functionalized graphene quantum dots (His-GQDs) form enantiomeric nanoscale structures with distinct interfacial topologies despite closely matched size, composition, and surface charge. D-His-GQDs preferentially interact with cholesterol-rich ordered membrane nanodomains associated with S-acylated coronavirus Spike, penetrate more deeply into raft-like lipid bilayers, and induce lipid disordering, membrane leakage, and viral-envelope disruption. Reducing Spike S-acylation or depleting membrane cholesterol attenuated this stereoselective interaction, while molecular simulations revealed asymmetric lipid organization surrounding S-acylated Spike and preferential insertion of D-His-GQDs into the ordered membrane environment. Functionally, D-His-GQDs directly inactivated human coronavirus OC43 (HCoV-OC43), inhibited infection with an EC50 of 0.92 g/mL and exhibited a selectivity index of 361. Stereoselective antiviral activity extended to human cytomegalovirus, demonstrating activity across distinct enveloped-virus families, and intranasal D-His-GQDs protected HCoV-OC43-challenged mice under both prophylactic and early post-exposure regimens. Overall, these findings establish structural nanochirality as a biomaterial design parameter for recognizing higher-order membrane organization and converting virus-associated membrane states into selectively addressable therapeutic vulnerabilities.

The blood-brain barrier (BBB) constitutes a major bottleneck for the systemic delivery of most therapeutic agents to the central nervous system. Here, we report near-infrared reversible optoacoustic modulation of the BBB permeability (NIR-ROAMBBB), leveraging endothelial tight junction targeting hyperbranched gold nanoconstructs (HBGNCs) to amplify localized optoacoustic transduction under femtosecond laser excitation. We first synthesized HBGNCs with tunable particle sizes (62-150 nm) and consistent branch morphologies via a seed-mediated growth approach, and uncovered a non-monotonic relationship between particle dimension and optoacoustic output, where the 62 nm HBGNCs generated nearly twofold stronger optoacoustic signal than gold nanorods and gold nanostars under matched excitations. Conjugation with BV11 antibodies against junctional adhesion molecule A increased HBGNC endothelial association and cerebral accumulation, enabling focal and fluence-dependent transient BBB opening (3-6 h) under 800 nm femtosecond pulsed laser excitation, as validated by in vitro trans-endothelial electrical resistance measurements, ex vivo Evans blue extravasation staining, and in vivo NIR imaging. Featuring deep tissue penetration of NIR light, robust optoacoustic conversion of HBGNCs, and negligible femtosecond laser-induced photothermal damage, this non-invasive strategy enables precise focal modulation of BBB permeability and potential drug delivery.

Existing biological nanopores are often too narrow to accommodate large proteins and biomolecular complexes in their native folded states, precluding the analysis of megadalton assemblies. Here, we report the self-assembly of the cholesterol-dependent cytolysin Perfringolysin O (PFO) into stable transmembrane nanopores composed of 46 {+/-} 9 monomers, with an inner pore diameter of 28.5 {+/-} 5.6 nm and a length of 9.8 nm. We demonstrate that despite the large size of the pore assembly, PFO pores exhibit a stable open-pore current with a high signal-to-noise ratio, making them suitable for resistive-pulse recordings. Moreover, PFO nanopores enable accurate, calibration-free, and label-free sizing of individual proteins, multi-protein complexes, and viral particles across a large molecular-weight range spanning 50 kDa to 3.4 MDa. The exceptionally large diameter of these pores enables, for the first time, resistive-pulse-based characterization of intact virus particles and ribosomes with a biological nanopore. Specifically, we determined the volume, shape, and diameter of complete capsids of recombinant adeno-associated virus serotype 2 (rAAV2) as well as capsid fragments. Finally, simultaneous analysis of molecular volume and shape resolved intact 70S ribosomes from their dissociated 30S and 50S subunits in a mixture. By extending biological nanopore sensing to single-particle characterization of large protein complexes well beyond the reach of existing pores, this approach introduces PFO nanopores as a versatile platform for label-free, single-particle analysis in solution.