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Cryo-electron tomography with subtomogram averaging is useful for the structural analysis of heterogeneous protein complexes. emClarity is graphics processing unit-accelerated image-processing software for subtomogram averaging and classification at high resolution.
In this protocol, CRISPR perturbations are introduced in cells using lentiviral libraries and read out using in situ sequencing, coupling high-throughput pooled screening with phenotypic image readouts in live or fixed cells.
The authors describe how to establish dissociated cultures of neurons, astrocytes, microglia, pericytes, and brain endothelial and neural precursor cells, as well as explant cultures of the leptomeninges, cortical slice cultures and brain tumor cells.
IBEX (iterative bleaching extends multiplexity) is an iterative immunolabeling and chemical bleaching method that enables highly multiplexed imaging in diverse tissues.
KAS-seq is an N3-kethoxal–assisted single-stranded DNA sequencing approach that allows rapid, sensitive and genome-wide mapping of single-stranded DNA produced in situ using as few as 1,000 cells or animal tissues.
This protocol describes an experimental and computational approach for mapping higher-order DNA interactions that relies on tagging cross-linked fragmented chromatin through an iterative split-and-pool barcoding process.
Star-shaped DNA nanostructures are designed to inhibit dengue virus in vitro. The ‘DNA star’ acts as a template to display ten virus-binding aptamers with precise spatial patterning to mirror the virus antigens, achieving high viral binding avidity.
Bacterial genome-wide gene fitness is assessed by CRISPRi-seq. The procedure includes a pipeline for single-guide RNA library design, workflows for pooled CRISPRi library construction, growth assays, sequencing and read analysis fitness quantification.
This protocol describes an in vitro model of the human blood–brain barrier, self-assembled within microfluidic devices from stem-cell-derived or primary brain endothelial cells, and primary brain pericytes and astrocytes.
A protocol is described for generating human brain assembloids and performing viral labeling and retrograde tracing, 3D live imaging of axon projection and optogenetics with calcium imaging and electrophysiological recordings to model neural circuits.