Nature Protocols

Article	Citations
Tutorial: a guide to performing polygenic risk score analyses	979
Using MetaboAnalyst 5.0 for LC–HRMS spectra processing, multi-omics integration and covariate adjustment of global metabolomics data	697
The trRosetta server for fast and accurate protein structure prediction	334
Establishment of patient-derived cancer organoids for drug-screening applications	332
Quantification of SARS-CoV-2 neutralizing antibody by a pseudotyped virus-based assay	307
CODEX multiplexed tissue imaging with DNA-conjugated antibodies	275
Tutorial: design and fabrication of nanoparticle-based lateral-flow immunoassays	267
A practical guide to large-scale docking	240
Design, fabrication and applications of tetrahedral DNA nanostructure-based multifunctional complexes in drug delivery and biomedical treatment	232
Isolation and characterization of extracellular vesicle subpopulations from tissues	221
Quantification of SARS-CoV-2 neutralizing antibody by wild-type plaque reduction neutralization, microneutralization and pseudotyped virus neutralization assays	208
Minimum Information for Reporting on the Comet Assay (MIRCA): recommendations for describing comet assay procedures and results	205
Efficient low-cost chromatin profiling with CUT&Tag	201
Proximity labeling in mammalian cells with TurboID and split-TurboID	193
Tutorial: guidelines for the computational analysis of single-cell RNA sequencing data	187
Metagenome analysis using the Kraken software suite	172
Global analysis of repetitive DNA from unassembled sequence reads using RepeatExplorer2	159
Tutorial: guidelines for standardized performance tests for electrodes intended for neural interfaces and bioelectronics	159
MOF-derived nanoporous carbons with diverse tunable nanoarchitectures	151
Artificial intelligence–enabled virtual screening of ultra-large chemical libraries with deep docking	145
I-TASSER-MTD: a deep-learning-based platform for multi-domain protein structure and function prediction	144
Using single-vesicle technologies to unravel the heterogeneity of extracellular vesicles	141
Engineering SARS-CoV-2 using a reverse genetic system	140
Generation and long-term culture of advanced cerebral organoids for studying later stages of neural development	137
Chromatin accessibility profiling by ATAC-seq	135

Preparation of selective organ-targeting (SORT) lipid nanoparticles (LNPs) using multiple technical methods for tissue-specific mRNA delivery	133
Tutorial: best practices and considerations for mass-spectrometry-based protein biomarker discovery and validation	125
Optimization of metabolomic data processing using NOREVA	119
Multiplexed single-cell proteomics using SCoPE2	119
High-yield production of mono- or few-layer transition metal dichalcogenide nanosheets by an electrochemical lithium ion intercalation-based exfoliation method	119
3D bioprinted silk fibroin hydrogels for tissue engineering	118
Standardized bacteriophage purification for personalized phage therapy	117
Long-term culture, genetic manipulation and xenotransplantation of human normal and breast cancer organoids	113
Tutorial: guidelines for annotating single-cell transcriptomic maps using automated and manual methods	112
Chemometric analysis in Raman spectroscopy from experimental design to machine learning–based modeling	109
An in vitro batch fermentation protocol for studying the contribution of food to gut microbiota composition and functionality	108
Manual and automated preparation of single-stranded DNA libraries for the sequencing of DNA from ancient biological remains and other sources of highly degraded DNA	107
Algorithmic assessment of cellular senescence in experimental and clinical specimens	104
Intestinal organoid cocultures with microbes	104
High-yield, green and scalable methods for producing MOF-303 for water harvesting from desert air	103
Measuring DNA modifications with the comet assay: a compendium of protocols	102
High-throughput cultivation and identification of bacteria from the plant root microbiota	101
Ligand binding free-energy calculations with funnel metadynamics	100
Precision genome editing using cytosine and adenine base editors in mammalian cells	99
Jointly defining cell types from multiple single-cell datasets using LIGER	97
A standardized social preference protocol for measuring social deficits in mouse models of autism	97
Generation and biobanking of patient-derived glioblastoma organoids and their application in CAR T cell testing	96
Controlling the polarity of human gastrointestinal organoids to investigate epithelial biology and infectious diseases	96
RNA-extraction-free nano-amplified colorimetric test for point-of-care clinical diagnosis of COVID-19	95
Engineered human blood–brain barrier microfluidic model for vascular permeability analyses	95
Microfluidic integration of regeneratable electrochemical affinity-based biosensors for continual monitoring of organ-on-a-chip devices	94
Establishment and differentiation of long-term trophoblast organoid cultures from the human placenta	91
Generation of insulin-producing pancreatic β cells from multiple human stem cell lines	89
Analyzing high-dimensional cytometry data using FlowSOM	87
Reliable detection of somatic mutations in solid tissues by laser-capture microdissection and low-input DNA sequencing	87
Rolling circle amplification (RCA)-based DNA hydrogel	86
A dynamic DNA tetrahedron framework for active targeting	85
3D in vitro morphogenesis of human intestinal epithelium in a gut-on-a-chip or a hybrid chip with a cell culture insert	82
Establishment of human fetal hepatocyte organoids and CRISPR–Cas9-based gene knockin and knockout in organoid cultures from human liver	80
A metabolic modeling platform for the computation of microbial ecosystems in time and space (COMETS)	78
Dynamic single-cell phenotyping of immune cells using the microfluidic platform DropMap	78
Combined proximity labeling and affinity purification−mass spectrometry workflow for mapping and visualizing protein interaction networks	75
Identifying unknown metabolites using NMR-based metabolic profiling techniques	73
The normative modeling framework for computational psychiatry	73
Engineering brain assembloids to interrogate human neural circuits	73
Bioconjugates of photon-upconversion nanoparticles for cancer biomarker detection and imaging	71
Microtiter plate assays to assess antibiofilm activity against bacteria	71
lentiMPRA and MPRAflow for high-throughput functional characterization of gene regulatory elements	70
Measuring the elastic modulus of soft culture surfaces and three-dimensional hydrogels using atomic force microscopy	70
Generic acquisition protocol for quantitative MRI of the spinal cord	69
Fabrication, characterization and applications of graphene electronic tattoos	67
Improved high-molecular-weight DNA extraction, nanopore sequencing and metagenomic assembly from the human gut microbiome	66
Detection of aberrant gene expression events in RNA sequencing data	64
Antigen retrieval and clearing for whole-organ immunofluorescence by FLASH	64
Genetic circuit design automation with Cello 2.0	63

A robust unsupervised machine-learning method to quantify the morphological heterogeneity of cells and nuclei	63
Accurate determination of protein:ligand standard binding free energies from molecular dynamics simulations	62
Tools for experimental and computational analyses of off-target editing by programmable nucleases	61
Whole-brain functional ultrasound imaging in awake head-fixed mice	60
NovoSpaRc: flexible spatial reconstruction of single-cell gene expression with optimal transport	60
Tutorial: practical considerations for tissue clearing and imaging	58
Nanocarriers based on bacterial membrane materials for cancer vaccine delivery	58
Digital postprocessing and image segmentation for objective analysis of colorimetric reactions	58
Rapid (30-second), equipment-free purification of nucleic acids using easy-to-make dipsticks	58
Synthesis of siRNA nanoparticles to silence plaque-destabilizing gene in atherosclerotic lesional macrophages	58
Solid-state 31P NMR mapping of active centers and relevant spatial correlations in solid acid catalysts	57
A complete pupillometry toolbox for real-time monitoring of locus coeruleus activity in rodents	57
A well plate–based multiplexed platform for incorporation of organoids into an organ-on-a-chip system with a perfusable vasculature	57
Comparative analysis of assays to measure CAR T-cell-mediated cytotoxicity	56
Cargo loading within ferritin nanocages in preparation for tumor-targeted delivery	56
CRISPR–Cas9, CRISPRi and CRISPR-BEST-mediated genetic manipulation in streptomycetes	56
Generation, functional analysis and applications of isogenic three-dimensional self-aggregating cardiac microtissues from human pluripotent stem cells	56
Norovirus detection in water samples at the level of single virus copies per microliter using a smartphone-based fluorescence microscope	55
Synthesis of N-heterocyclic carbene gold(I) complexes	55
Tutorial: avoiding and correcting sample-induced spherical aberration artifacts in 3D fluorescence microscopy	55
Tutorial: using nanoneedles for intracellular delivery	54
Raman spectroscopy for real-time and in situ monitoring of mechanochemical milling reactions	53
Stem-cell-derived human microglia transplanted into mouse brain to study human disease	53
Physical dynamic double-network hydrogels as dressings to facilitate tissue repair	53
Combined whole-mount fluorescence in situ hybridization and antibody staining in zebrafish embryos and larvae	52
Analysis of clonogenic growth in vitro	52
A complete and flexible workflow for metaproteomics data analysis based on MetaProteomeAnalyzer and Prophane	51
Transplantation of intestinal organoids into a mouse model of colitis	50
Stable isotope tracing to assess tumor metabolism in vivo	50
Comprehensive structural glycomic characterization of the glycocalyxes of cells and tissues	50
A universal approach for the synthesis of mesoporous gold, palladium and platinum films for applications in electrocatalysis	49
Mapping mechanical properties of biological materials via an add-on Brillouin module to confocal microscopes	48
Evaluation of the effects of cold atmospheric plasma and plasma-treated liquids in cancer cell cultures	48
Culture and analysis of kidney tubuloids and perfused tubuloid cells-on-a-chip	48
Creating CRISPR-responsive smart materials for diagnostics and programmable cargo release	48
Characterization of connective tissues using near-infrared spectroscopy and imaging	47
Quantitative nucleotide resolution profiling of RNA cytidine acetylation by ac4C-seq	47
Inducible mouse models of colon cancer for the analysis of sporadic and inflammation-driven tumor progression and lymph node metastasis	47
Designing and executing prime editing experiments in mammalian cells	47
Optimized single-nucleus transcriptional profiling by combinatorial indexing	47
Phage-assisted continuous and non-continuous evolution	46
Use of deep learning to develop continuous-risk models for adverse event prediction from electronic health records	46
Design and simulation of DNA, RNA and hybrid protein–nucleic acid nanostructures with oxView	46
Tutorial: a computational framework for the design and optimization of peripheral neural interfaces	45
IBEX: an iterative immunolabeling and chemical bleaching method for high-content imaging of diverse tissues	45
Isolation of mitochondria-derived mitovesicles and subpopulations of microvesicles and exosomes from brain tissues	45
High-resolution in situ structure determination by cryo-electron tomography and subtomogram averaging using emClarity	44
In vitro generation of self-renewing human intestinal epithelia over planar and shaped collagen hydrogels	44
Design, assembly, and characterization of membrane-spanning DNA nanopores	44
Comprehensive isolation of extracellular vesicles and nanoparticles	43
Statistical analysis in metabolic phenotyping	43
Monomicellar assembly to synthesize structured and functional mesoporous carbonaceous nanomaterials	42
Optofluidic Raman-activated cell sorting for targeted genome retrieval or cultivation of microbial cells with specific functions	42
A generalized workflow for conducting electric field–optimized, fMRI-guided, transcranial magnetic stimulation	41
Spooling electrochemiluminescence spectroscopy: development, applications and beyond	41
Composable microfluidic spinning platforms for facile production of biomimetic perfusable hydrogel microtubes	40
Use of human peripheral blood mononuclear cells to define immunological properties of nucleic acid nanoparticles	40
Continuous human iPSC-macrophage mass production by suspension culture in stirred tank bioreactors	40
Complex-centric proteome profiling by SEC-SWATH-MS for the parallel detection of hundreds of protein complexes	40
Preparation and quantitative analysis of multicenter luminescence materials for sensing function	40
Chronic, cortex-wide imaging of specific cell populations during behavior	40
A plate-based single-cell ATAC-seq workflow for fast and robust profiling of chromatin accessibility	40
A comprehensive guide to studying inflammasome activation and cell death	38
Implantation of Neuropixels probes for chronic recording of neuronal activity in freely behaving mice and rats	38
All-optical interrogation of neural circuits in behaving mice	38
Organotypic culture assays for murine and human primary and metastatic-site tumors	38
Engineering DNA nanostructures for siRNA delivery in plants	38
Coculture of primary human colon monolayer with human gut bacteria	37
Spatially resolved analysis of FFPE tissue proteomes by quantitative mass spectrometry	37
A quantitative thiol reactivity profiling platform to analyze redox and electrophile reactive cysteine proteomes	37
Quantification of organelle contact sites by split-GFP-based contact site sensors (SPLICS) in living cells	36
Testing physiologically relevant conditions in minimal inhibitory concentration assays	36
Smart-RRBS for single-cell methylome and transcriptome analysis	36
Tutorial: assessing metagenomics software with the CAMI benchmarking toolkit	36
An optimized comet-based in vitro DNA repair assay to assess base and nucleotide excision repair activity	36
Creating hierarchical pores in metal–organic frameworks via postsynthetic reactions	36
The maternal serum metabolome by multisegment injection-capillary electrophoresis-mass spectrometry: a high-throughput platform and standardized data workflow for large-scale epidemiological studies	35
Expression and characterization of SARS-CoV-2 spike proteins	35
Perfusion and endothelialization of engineered tissues with patterned vascular networks	35
Protein higher-order-structure determination by fast photochemical oxidation of proteins and mass spectrometry analysis	35

Seed amplification assay for the detection of pathologic alpha-synuclein aggregates in cerebrospinal fluid	34
Responsive and activable nanomedicines for remodeling the tumor microenvironment	34
iBEAT V2.0: a multisite-applicable, deep learning-based pipeline for infant cerebral cortical surface reconstruction	34
CRISPRi-seq for genome-wide fitness quantification in bacteria	34
Tutorial: structural characterization of isolated metal atoms and subnanometric metal clusters in zeolites	34
Defining genome-wide CRISPR–Cas genome-editing nuclease activity with GUIDE-seq	34
Integrative structural modeling of macromolecular complexes using Assembline	34
High-throughput matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) mass spectrometry–based deubiquitylating enzyme assay for drug discovery	34
Generation and characterization of hair-bearing skin organoids from human pluripotent stem cells	34
Transcriptome-wide identification of RNA-binding protein binding sites using seCLIP-seq	33
Graphene- and metal-induced energy transfer for single-molecule imaging and live-cell nanoscopy with (sub)-nanometer axial resolution	33
Single-particle tracking photoactivated localization microscopy of membrane proteins in living plant tissues	33
Label-free capacitive assaying of biomarkers for molecular diagnostics	33
Imaging and optogenetic modulation of vascular mural cells in the live brain	33