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This protocol describes how to undertake high spatial and temporal Ca2+ imaging of ex vivo multicellular myocardial strips that include the endocardial surface, allowing the study of the Ca2+ signaling that underpins cardiomyocyte contraction.
This protocol comprises various methods to coculture organoids (particularly human small intestinal and colon organoids) with microbes, including microinjection into the lumen and periphery of 3D organoids and exposure of organoids to microbes in a 2D layer.
This protocol outlines the experimental and computational steps of MetaRibo-Seq, a modified ribosome profiling approach that allows direct interrogation of translation in uncultured bacterial communities.
This protocol describes a complete workflow for detecting significant contacts in Capture Hi-C data, including preprocessing, interaction calling and downstream analyses, based on the CHiCAGO pipeline and companion tools.
The authors provide a protocol for fluorescence multiplex host cell reactivation, which uses reporter plasmids containing site-specific DNA lesions to assess DNA repair capacity in at least six major DNA repair pathways in live cells.
Radiotracers used in human positron emission tomography studies need to be prepared according to current good manufacturing practice. Here we use the synthesis of [11C]ER176 as an example to show the translation of an experimental radiosynthesis to a current good manufacturing practice process.
Human pluripotent stem cells acquire cancer-related mutations during culture, which can impact their use in the clinic or in basic research. This protocol describes a computational pipeline to detect such mutations using RNA sequencing data.
The authors describe procedures for genome-wide quantification of histone modifications and chromatin-associated proteins after replication (chromatin occupancy after replication-seq) and for profiling their relative occupancy in nascent sister chromatids (sister chromatids after replication-seq).
This protocol describes how to rapidly isolate a specific cell type from mouse blood or tissues for mass-spectrometry-based metabolomics. Mice are infused with isotopically labeled compounds, and cells are isolated using antibody-coated magnetic beads.
Millman and colleagues describe a six-stage monolayer culture differentiation protocol for generating insulin-secreting pancreatic β cells from a variety of human pluripotent stem cell lines and outline steps for in vitro functional assessment.
This protocol describes a high-throughput sequencing approach to identifying regulators of bacterial gene expression based on transposon insertion mutagenesis in a strain harboring a fluorescent reporter.
This protocol describes novoSpaRc, a computational pipeline for de novo reconstruction of spatial gene expression from single-cell RNA sequencing with the potential to incorporate spatial atlas data to improve the reconstruction.
This high-throughput protocol for direct ex vivo real-time metabolic fingerprinting of biofluids uses a laser system and an automated sampling platform. It includes REIMS procedures for analyzing blood, urine, stool, saliva, sputum and breast milk.
This protocol describes how to use free-space angular-chirp-enhanced delay (FACED), an all-optical, passive and reconfigurable laser-scanning approach, to increase the imaging speed of various microscopy modalities.
This protocol describes isolation and long-read sequencing (using either the Oxford Nanopore or PacBio platforms) of nascent chromatin-associated RNAs from Arabidopsis seedlings and bioinformatic pipelines for profiling splicing and polyadenylation.
This protocol enables explicit regional and functional division of functional elements of solid-state nanochannels to improve the sensitivity and specificity of bioanalysis in complex matrices.
Metabolomics studies using large-scale NMR or mass spectrometry experiments on biofluids or tissues generate complex data. This protocol provides guidelines and software (supplied in Jupyter notebooks) for the statistical analysis of these data.
The glomerulus is a difficult-to-isolate structure that is often poorly represented in single-cell kidney preparations. This protocol describes the isolation of high-quality mouse glomerular cells for high-throughput analyses.
This protocol describes a plate-based ATAC-seq assay that combines up-front bulk tagging of accessible DNA by the Tn5 transposase with FACS sorting for robust and cost-efficient profiling of chromatin accessibility in single cells.