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Adult endothelial cells from mice are reprogrammed to hematopoietic stem cells without transitioning through a pluripotent state. The protocol also describes in vitro and in vivo assays to confirm that the resulting cells function as expected.
This protocol describes the differentiation of hiPSCs into cardiomyocytes and subsequent cytotoxicity and contractility assays needed to calculate the ‘cardiac safety index’, which models the likelihood that a drug is cardiotoxic.
This protocol provides approaches for applying the CRISPR–Cas9 system for genome editing in apple and grapevine plants, using both plasmid-mediated delivery of components and direct delivery of CRISPR–Cas9 ribonucleoproteins.
This computational protocol offers a framework to integrate high-dimensional -omics datasets. A three-pronged association study integrating intestinal microbiome and serum metabolome data with measures of human host physiology is used as an example.
Valuable structural information can be derived by chemical cross-linking of proteins followed by mass spectrometry of the products. Iacobucci et al. use MS-cleavable reagents, enrichment by strong-cation-exchange chromatography, and LC-MS/MS analysis.
Endothelial cells, pericytes and astrocytes are cocultured to generate organoids that reproduce many features of the blood–brain barrier. This protocol also describes how to analyze drug penetration into the organoids.
This protocol describes how to implement and apply an adaptive light-sheet microscopy framework (AutoPilot). The procedure can be used to introduce AutoPilot in an existing microscope or to set up a new adaptive multiview light-sheet microscope.
This protocol describes the synthesis, characterization, and calibration of a nanoscale fiber-optic force sensor. The sensor can be used to detect sub-piconewton forces and acoustic waves in biological environments by means of optical readouts.
This protocol describes how to generate transgenic zebrafish expressing a barcode array that can be edited by CRISPR–Cas9 at multiple developmental stages. Single-cell RNA sequencing of edited barcodes and cellular transcriptomes allows reconstruction of lineage relationships.
This protocol describes how to produce cell-laden microfibers using capillary microfluidic devices. The devices enable spinning of increasingly complex microfibers, which can function as building blocks for 3D cell culture and tissue engineering.
This protocol addresses the need to define informative priors to apply ensemble modeling in systems biology. The protocol collects parameters, assesses their plausibility and creates log-normal probability distributions for use as informative priors.
Spatial Transcriptomics combines histological staining and spatially resolved RNA-sequencing data from tissue sections. This protocol describes how to implement this method with mammalian tissue.
This protocol describes the synthesis of magnetic resonance tuning (MRET) sensors. The sensors consist of two magnetic components separated by a linker and can be modularly designed for targets such as enzymes, nucleic acid sequences, and pH values.
This protocol provides a procedure by which Drosophila ovarioles are dissected with or without the epithelial sheath and placed in a closed chamber that mimics physiological conditions for imaging.
This protocol describes CIRCLE-seq (circularization for in vitro reporting of cleavage effects by sequencing), a sensitive and unbiased method for defining the on-target and off-target activity of CRISPR–Cas9 nucleases genome-wide.
Hydroxyl-radical footprinting provides a wealth of data on the structure of nucleic acid–protein complexes. HYDROID is a software tool used to quantify footprinting data from gel electrophoresis images and integrate them with structural models.
Here, the authors describe how to use BasePlayer, an interactive and user-friendly software that facilitates the identification of causative mutations from next-generation sequencing data.
The immunoassay multiplexing capacity of single-cell mass cytometry relies on monoclonal antibodies labeled with stable heavy-metal isotopes. Nolan et al. describe procedures for conjugating monoclonal IgG antibodies with 48 heavy-metal isotopes.
This protocol describes how to combine up to four genetically encoded fluorescent sensors to image redox landscapes. The procedure describes applications in live imaging and flow cytometry of cultured cells, and in vivo imaging in zebrafish larvae.
This protocol describes an approach for pulse labeling of ventricular zone progenitors and their neuronal progeny in the developing brain. Labeled cells can be isolated and highlighted by FACS or immunohistochemistry.