Imaging mass cytometry (IMC) is a highly multiplexed tissue-imaging technology that enables the simultaneous detection of up to 45 markers in situ at subcellular resolution. By combining probes such as antibodies conjugated to metal isotopes with laser ablation and time-of-flight mass spectrometry, IMC generates spatially resolved, high-dimensional single-cell data. Since its introduction in 2014, IMC has become a widely adopted platform in spatial biology. Researchers have used IMC to advance our molecular-level understanding of cancer as well as autoimmune and infectious diseases, and the platform is also increasingly used in the translational and clinical setting. In this Primer, we provide a comprehensive guide to the experimental IMC workflow, covering antibody conjugation and panel design, sample preparation, data acquisition and quality control. We detail the computational analysis pipeline, including image preprocessing, segmentation, feature extraction, and both single-cell and spatial analyses. Key applications to biological and clinical research are highlighted. We discuss current limitations and optimization strategies. Finally, future directions are outlined, including IMC foundation models, multi-modal integration and the path to clinical implementation. This Primer seeks to provide both new users and IMC experts with detailed insights and practical guidance to unlock the full potential of IMC. Imaging mass cytometry is a highly multiplexed tissue-imaging technique that allows the simultaneous in situ detection of up to 45 markers with subcellular-level resolution. In this Primer, Meyer et al. provide a comprehensive guide to the experimental imaging mass cytometry workflow.
Positrons are antiparticles of electrons that can serve as ultrasensitive probes of atomic-level structural detail in materials. When positrons annihilate with electrons, characteristic γ-rays are produced. Positron annihilation spectroscopy is based on the detection of the γ-photons emitted in the positron–electron annihilation process, and experimental methods analyse the time, energy and angular distributions of these γ-photons. In crystalline solids, the selective sensitivity of positrons to vacancy defects allows one to identify their structure and to determine their concentrations and physical properties in a wide range of materials. This Primer presents techniques in positron annihilation spectroscopy, evaluates the typical results obtained and highlights recent advances across various fields, ranging from point defects and porosity in solids to material surfaces and interfaces. This Primer provides an introduction to positron annihilation spectroscopy and its uses in studying atomic-scale structures in solids.
Detection sensitivity fundamentally limits imaging depth and signal-to-noise ratio in photoacoustic tomography (PAT). This Primer summarizes the noise-equivalent pressures (NEP) reported in the literature for piezoceramic, piezopolymer, capacitive micromachined ultrasonic transducer (CMUT) and optical ultrasound detectors applicable to PAT as a function of detector size and bandwidth. Millimetre-scale piezoceramic transducers achieve NEPs as low as 0.06 mPa/√Hz, enabling the detection of sub-pascal acoustic pressures at low-megahertz frequencies. Meanwhile, sub-100-µm-sized optical ultrasound sensors can provide NEPs of 0.6 mPa/√Hz, enabling the detection of a few pascals at frequencies in the tens of megahertz range. Piezopolymer detectors, meanwhile, exhibit higher NEPs but with broader bandwidths, whereas CMUTs provide NEPs comparable to the lowest available for 0.1–1-mm-sized detectors, albeit with resonant responses. By establishing and explaining the NEP landscape, this Primer aims to aid the understanding and selection of detectors for PAT and inform their future development. Photoacoustic tomography (PAT) is a biomedical imaging technique based on the use of optically excited ultrasound waves. In this Primer, Guggenheim et al. describe the noise-equivalent pressures reported in the literature for detectors applicable to PAT.
Neutron spin echo (NSE) spectroscopy is a powerful technique used to probe the internal dynamics and diffusion processes within matter, allowing researchers to quantify how materials respond to changes in external conditions. To fully understand the bulk properties of a material, it is often crucial to investigate dynamic processes occurring on nanometre length scales and nanosecond timescales. NSE spectroscopy provides an exceptional capability to measure such dynamics across a broad spectrum of condensed matter systems, ranging from proteins and quantum magnets to catalysts. In this Primer, we present an overview of NSE spectroscopy, highlighting the instruments used, methods of data collection and representative applications in both soft-matter and hard-matter sciences. Neutron spin echo spectroscopy investigates internal dynamics and diffusion in materials, using advanced instruments and data collection techniques. In this Primer, Faraone et al. discuss how neutron spin echo spectroscopy can be used in both soft-matter and hard-matter sciences.
G protein-coupled receptors (GPCRs) represent the largest class of therapeutic targets, and peptides have become an increasingly important and versatile ligand type for studying receptor biology and developing new drugs. Although many core pharmacological concepts apply to all ligand classes, recent advances in peptide-focused approaches, from innovative discovery strategies and combinatorial library synthesis to computational design and structural integration, have substantially broadened the GPCR drug discovery toolbox. In this Primer, we outline experimental and computational workflows tailored to peptide–GPCR interactions, including in silico peptide mining, deorphanization strategies, library-based screening platforms, modern pathway-resolved biosensor assays, and approaches for peptide stabilization and optimization strategies to address their pharmacokinetic limitations. We summarize recent progress in structural modelling, diffusion-based de novo design, molecular dynamics simulations, and free energy calculations, as well as artificial intelligence-guided or machine learning-guided screening frameworks that connect peptide sequence space with receptor binding and functional signalling outcomes. We also discuss key challenges in the field, including reproducibility issues, ambiguity in sequence annotation and post-translational modifications, peptide instability, assay artefacts and current limitations of computational mining approaches, and we propose practical strategies to address them. Finally, we outline future perspectives, emphasizing integrated workflows that combine experimental pharmacology, structural biology and computational modelling to accelerate the discovery of next-generation peptide probes and therapeutic ligands targeting GPCRs. Peptides have become an increasingly important and versatile ligand type for G protein-coupled receptors (GPCRs). In this Primer, Hermes et al. discuss the experimental and computational workflows for assessing peptide–GPCR interactions.
Conductive polymers (CPs) are π-conjugated organic semiconductors that exhibit electrical conductivities spanning those of insulators to metals, depending on the doping level, while retaining the mechanical properties of plastics. This combination positions CPs as a promising class of advanced functional materials. CPs hold several distinctive properties, including solution processability, tunable optoelectronic properties, mechanical ductility and biocompatibility, making them suitable for next-generation optoelectronic and bioelectronic devices. This Primer provides an overview of the conduction and doping mechanisms of CPs and discusses typical synthetic methods used for their preparation. It also highlights key structural, macromolecular, optical and charge transport properties that govern their functionalities and device performance. Finally, representative applications in flexible electronic circuits, biosensors and tissue engineering are discussed, emphasizing the potential of CPs to revolutionize commercial electronics, energy conversion and healthcare. Despite several challenges and performance limitations, recent advancements in synthetic methodologies, doping strategies and device engineering are driving the commercialization of CP-based technologies. Conductive polymers (CPs) are π-conjugated organic semiconductors that exhibit electrical conductivities similar to insulators and metals while maintaining the flexibility of plastics. This Primer provides an overview of the synthesis, conduction and doping mechanisms of CPs while emphasizing their functionality in flexible electronic circuits, biosensors and tissue engineering.
Matrix-assisted laser desorption/ionization imaging mass spectrometry (MALDI IMS) is a transformative molecular imaging technology capable of mapping diverse chemical classes, from small metabolites, neurotransmitters and lipids to N-glycans and proteins, at cellular resolution. The field has rapidly matured, offering a range of mass analysers, including axial time-of-flight, quadrupole (or orthogonal) time-of-flight, and high-resolution Orbitrap and Fourier-transform ion cyclotron resonance systems, each with distinct characteristics in terms of spatial resolution, chemical specificity and throughput. In practice, MALDI IMS involves applying a light-absorbing chemical matrix onto a tissue section, followed by automated laser irradiation at discrete coordinates (like pixels) to desorb and ionize endogenous molecules for mass analysis. This label-free approach preserves spatial context, providing a molecular map that links highly multiplexed molecular distributions to distinct anatomical regions, functional tissue units and cell types in situ. This Primer includes an overview of basic imaging MALDI IMS concepts, instrumentation, data processing approaches and advanced applications. We also address key challenges and considerations, with an eye towards optimizing instrumentation and methods to overcome issues in spatial resolution, sensitivity and specificity. Finally, we look towards the future of the technology, including its integration with other spatial omics modalities and its potential as a tool for precision medicine. Matrix-assisted laser desorption/ionization imaging mass spectrometry (MALDI IMS) is a label-free molecular imaging approach mapping diverse biomolecules at cellular resolution. In this Primer, Spraggins et al. outline MALDI IMS concepts, instrumentation, data processing and integration with spatial omics and medicine.
This Primer features cucurbit[n]uril(CB[n])-type receptors, a family of synthetic macrocycles that function as molecular containers for guest compounds in water, where n is the number of glycoluril units linked by methylene bridges to form a barrel-shaped cavity. Encapsulation within CB[n] hosts can markedly alter the physical, chemical, optical and biological properties of bound guests, enabling applications in chemical sensing, bioimaging, chemical biology, drug delivery and drug sequestration, separations, catalysis and functional materials. In this Primer, we provide an overview of the synthesis of macrocyclic CB[n], functionalized CB[n] and acyclic CB[n]-type receptors. The defining feature of CB[n] hosts is their exceptionally high binding affinity (Ka, routinely >106 M−1) for hydrophobic cations in buffered aqueous media, which arises primarily from the release of high-energy waters from the host cavity upon complexation. Whereas CB[6] and CB[7] can form binary (host:guest, 1:1) complexes, CB[8] can form both binary and ternary (1:2) complexes featuring guest homodimerization or heterodimerization. CB[n]•guest complexes, where • indicates a supramolecular complex between the CB[n] macrocycle and a guest molecule, are responsive to chemical, photochemical, pH and electrochemical stimuli, enabling reversible switching or actuation of their bound state. Together, this Primer connects synthesis, molecular recognition mechanisms and functional outcomes in cucurbit[n]uril-type receptors. Cucurbit[n]urils are macrocyclic host molecules that form strong complexes with hydrophobic and cationic guests in water, where n is the number of glycoluril units. Cucurbit[n]urils complexed with guest molecules are stimuli-responsive, making them useful in applications such as functional materials, chemical sensing and separations, drug delivery systems and bioimaging.
Genome-wide mapping of factors that bind, package, replicate and transcribe DNA continues to be challenging despite rapid advances in DNA sequencing technologies. For nearly two decades, chromatin immunoprecipitation with sequencing (ChIP–seq) has dominated epigenomic mapping applications; however, alternative methods for epigenomic mapping have recently begun to replace ChIP–seq. The most popular alternatives use enzymes that modify, cleave or transpose DNA, using freely diffusing enzymes for chromatin accessibility mapping, or tethered ones to specific chromatin proteins or modifications for epigenomic profiling. Unlike ChIP–seq, which solubilizes chromatin before immunoprecipitation, enzyme-tethering epigenomic methods maintain cells or nuclei intact during DNA modification, cleavage or transposition. Although the basic in vivo and in situ enzyme-tethering technologies were introduced more than 20 years ago, they are especially well suited for current epigenomic challenges, including single-cell and spatial applications and both DNA-based and RNA-based time-resolved epigenomic mapping. In this Primer, we discuss DNA modification-based (DamID), cleavage-based (ChIC and CUT&RUN) and transposase-based (CUT&Tag and RT&Tag) tethered-enzyme profiling methods, and describe protocols for data processing, quality control and data analysis used by the epigenomics community. We anticipate that advances in enzyme engineering and automation will make enzyme-tethering methods increasingly attractive for both large and small epigenomic profiling projects. Enzyme-tethering epigenomic methods modify DNA while cells and nuclei remain intact, making them particularly suitable for single-cell and spatial applications. In this Primer, Kami Ahmad et al. discuss the various approaches for enzyme-tethering analysis and major applications in biological research.
Broadband spectroscopy combines wide spectral coverage with high resolution and accuracy, requirements that are often difficult to satisfy simultaneously using conventional approaches. Dual-comb spectroscopy addresses this challenge by using two optical frequency combs with slightly different repetition frequencies to map optical spectra directly into the radio-frequency domain. The method relies on time-domain interferometry and avoids mechanical scanning, enabling precise, rapid and broadband measurements. In principle, the achievable spectral resolution is set by the comb line spacing, defined by the laser repetition frequency. Because the measurement does not rely on geometrical constraints, the dual-comb interferometer offers a conceptual route towards broadband spectroscopy with resolution determined purely by temporal coherence. Over the past two decades, dual-comb spectroscopy has been implemented across the electromagnetic spectrum, from the terahertz to the visible, with ongoing efforts towards the ultraviolet. In this Primer, we present the fundamental principles of the technique, analyse the parameters governing its performance - including resolution, accuracy, signal-to-noise ratio and acquisition speed - and describe practical approaches to data acquisition and processing. We conclude by examining representative applications, current limitations and emerging directions for further development.
Flow nuclear magnetic resonance (NMR) spectroscopy is a powerful method for monitoring chemical reactions to improve their understanding and allow rational optimization. Flow NMR consists of making the reaction mixture flow through the NMR detector and can be tailored for the analysis of either batch or flow reactions. It provides quantitative measurements together with structural information over a broad range of conditions. The practice of flow NMR requires specialized hardware and know-how, some of which involves unusual spin effects, and requires customized methods for data collection and analysis. This Primer gives an overview of the features of flow NMR for chemical synthesis aimed at newcomers to the field. The reward of mastering flow NMR practice is illustrated through a selection of representative applications to mechanistic studies and reaction optimization in fields covering catalysis, polymer chemistry and autonomous synthesis. The range of flow NMR applications is continuously expanding thanks to new method developments that are described together with key challenges and limitations of the technique. Flow NMR spectroscopy offers real-time monitoring of chemical reactions, enabling quantitative analysis and mechanistic insights. This Primer describes the specialized equipment and tailored methods that can be applied to enhance its application across various fields, and addresses key challenges and limitations.
A laser Doppler vibrometer is an interferometric sensor with broadband demodulation of the photodetector signal used to measure movements, vibrations and deformations that occur in biological and technical systems. These time-dependent displacements can span many orders of magnitude in velocity, frequency bandwidth and amplitude. The velocity of movement can vary from zero to hundreds of metres per second, frequencies can range from microhertz to gigahertz and displacement amplitudes can occur down to the femtometre regime. Huge structures with kilometre-scale dimensions may be analysed, or structures may have micrometre or even nanometre dimensions. The immense breadth of scale and the potentially inaccessible or extremely sensitive nature of these motions make them difficult to measure with traditional sensors but can be probed with laser Doppler vibrometry. This Primer gives a comprehensive overview of laser Doppler vibrometry with different methods and applications presented. Although a wide diversity of sensing solutions exists, the field still offers many research opportunities, for example, gigahertz vibrometry with resolutions in the attometre regime using squeezed light on nanostructures, fast tracking of moving targets or measuring on rapidly moving or extremely sensitive surfaces. This Primer provides a comprehensive overview of laser Doppler vibrometry, covering fundamental principles, advanced measurement methods and diverse applications. It highlights performance limits, measurement ranges, future opportunities and practical considerations, including laser safety and calibration, and is supported by extensive supplementary information.
Current nanoherbicide development is largely guided by a material-first design logic that overlooks the biological complexity determining nanoparticle transport, transformation and target-site access in plants. We propose plant-informed nanodesign; a biology-driven framework that integrates plant physiology into rational nanocarrier engineering, advancing the field from empirical screening towards predictive science.
Base editing is a precision genome-editing methodology that enables the programmable installation of point mutations with high efficiency. Two major classes of base editors have been developed: cytosine base editors introducing C·G-to-T·A edits, and adenine base editors introducing A·T-to-G·C edits. Since their introduction, base editor use has expanded substantially, and base editors have been applied in many biological and biomedical applications. In this Primer, we provide an overview of the use of base editors in mammalian cells for high-throughput base-editing screens, disease modelling and therapeutic applications. We cover important considerations in experimental design of base-editing experiments, data analysis and interpretation, and best practices for reporting results. We also discuss potential challenges related to reproducibility and limitations, and outline options for optimization and troubleshooting. Our goal is to aid both new and experienced researchers in effectively implementing base editing in their own work. Base editing enables precise, programmable single-base changes using cytosine or adenine editors. This Primer outlines their use in mammalian cells for screening, disease modelling and therapy, covering experimental design, analysis, limitations and best-practice guidance to support effective implementation.
Magnetoresistance, the change of electrical resistance under a magnetic field, is one of the most versatile tools for probing charge, spin and orbital dynamics in solids. Since its discovery, magnetoresistance has provided unique insights into transport phenomena across classical, correlated and topological regimes, while also enabling technologies from magnetic sensors to data storage. This Primer introduces the physical principles that are fundamental to magnetoresistance and discusses some of the most important theoretical models that have been developed to account for them, ranging from Drude and Boltzmann transport to quantum linear response. Experimental methodologies to measure magnetoresistance in thin films and bulk materials are discussed, including device preparation, electrode design, measurement geometries and strategies to minimize artefacts such as current jetting, Hall mixing and parasitic capacitance. Finally, we discuss future opportunities in which magnetoresistance intersects with nanoscale fabrication, low dimensional materials and nonlinear transport, establishing it as a platform for both fundamental discoveries and emerging applications in quantum and functional devices. Magnetoresistance is a fundamental transport phenomenon that provides deep insight into electronic structure, magnetic order and emergent quantum effects in solids. This Primer reviews major magnetoresistance phenomena and presents practical measurement methodologies, emphasizing experimental configurations, instrumental limitations, common artefacts and best practices for reliable data interpretation.
Ultrafast pump–probe nano-imaging combines scanning probe-based optical near-field microscopy with ultrafast spectroscopy to enable imaging with deep sub-wavelength spatial resolution, femtosecond temporal resolution and simultaneous spectral resolution. Ultrafast nano-imaging has gained increased attention for its ability to provide far-from-equilibrium excitation and excited-state contrast. With coherent and nonlinear probing, coupled electron, spin and lattice dynamics on elementary timescale and length scale can be resolved. Through nano-movies, ultrafast nano-imaging visualizes correlated quantum dynamics underlying the properties of solid-state materials, semiconductors, molecular electronic, photonic, photovoltaic and other functional materials. With nanometre spatial resolution, this method probes elementary dynamic processes across multiple length scales that are otherwise obscured in conventional ultrafast spectroscopy in which heterogeneities are spatially averaged. This Primer describes the theoretical background and experimental implementation of ultrafast nano-imaging; signal interpretation and modelling; representative examples and a perspective for the future development of the field. Ultrafast near-field microscopy unites femtosecond optical spectroscopy with nanometre spatial resolution to image non-equilibrium material dynamics beyond the diffraction limit. This Primer outlines theory, instrumentation, signal interpretation and representative applications, providing a practical foundation for generating and analysing ultrafast nano-movies of quantum materials.
Quantum imaging encompasses many techniques, from the use of cameras that detect single photons with picosecond temporal resolution, to the use of nonlinear materials to create pairs of photons correlated over many different degrees of freedom, or even utilizing the interference between two of these photon-pair sources. This article focuses on the various imaging techniques using correlations between photon pairs, reviews the basic theory required to understand the method, discusses the practicalities of implementing these approaches, presents ways in which such systems might surpass classical limits and considers the applications that these advances might enable. Finally, we discuss future directions of the field of quantum imaging and the challenges that remain, both in terms of required new technologies and areas of physics in which our understanding may be incomplete. This Primer provides an introduction to quantum imaging with correlated photon pairs, from core concepts to experimental realizations. It discusses potential advantages over classical imaging, practical design issues, and the emerging applications and challenges that will shape future progress of the field.
Fluidic force microscopy is a versatile bionanotechnology platform that integrates atomic force microscopy with microfluidic probes. This hybrid approach enables precise measurement and application of forces across sub-nanonewton to micronewton ranges while simultaneously dispensing or sampling sub-femtolitre to picolitre volumes, all with real-time optical visualization at sub-micrometre resolution. In recent years, fluidic force microscopy has emerged as an enabling tool for minimally invasive single-cell manipulation, subcellular analysis and high-resolution nanoprinting applications. This Primer describes the fundamental principles underlying the combination of atomic force microscopy with microchannelled cantilevers, providing a comprehensive framework for understanding the unique capabilities of fluidic force microscopy and its rapidly expanding range of applications in biological research and nanotechnology. Fluidic force microscopy combines atomic force microscopy with microfluidic probes to enable measurement and manipulation of materials at sub-micrometre resolution. In this Primer, Zambelli et al. discuss the principles of fluidic force microscopy and applications in biological research and nanotechnology.
Electrophoretic deposition (EPD) is continuously evolving, transitioning from a method for applying macroscale coatings to a versatile platform for micropatterning and even single-particle manipulation. This evolution is driven by convergence with nanotechnology and precision manufacturing, enabling unprecedented control over the assembly of micromaterials and nanomaterials. This Primer elucidates the fundamental principles underpinning both macro-EPD and micro-EPD and discusses the interplay among suspension properties, kinetic parameters and deposition mechanisms that govern film composition, architecture and functionality. Innovative methodologies in electrode design and field modulation are highlighted, which achieve high-resolution patterning and multimaterial integration for emerging applications. The versatility of EPD renders it suitable for diverse applications, spanning protective coatings, energy, optoelectronics and biomedical areas. Finally, by analysing current limitations and optimization strategies, we offer insights into potential future directions for EPD development. Electrophoretic deposition is a micropatterning method that utilizes suspension properties and kinetic parameters for film control. This Primer discusses innovative electrode design to enhance resolution and multimaterial integration, addresses limitations and suggests future optimization strategies across diverse applications.
