Thank you for visiting nature.com. You are using a browser version with limited support for CSS. To obtain
the best experience, we recommend you use a more up to date browser (or turn off compatibility mode in
Internet Explorer). In the meantime, to ensure continued support, we are displaying the site without styles
and JavaScript.
Organoid culture systems are self-renewing 3D models derived from stem cells (pluripotent or adult-derived) or cancer cells that recapitulate characteristics of their tissues of origin. Here, we provide protocols for growing and studying various types of organoid (brain, skin, kidney, liver, bile duct, prostate, placenta, breast, colon, stomach, pancreas, lung, heart, and blood vessel) generated from both healthy and cancer cells.
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 summarizes the various approaches available to derive organoids from cancer patients and use these for screening of possible treatments. An optimized protocol for using head and neck cancer organoids is also described.
This multiplexed mass cytometry protocol uses thiol-reactive organoid barcoding in situ and a cytometry by time of flight signaling analysis pipeline (CyGNAL) to enable 126-plex single-cell analysis of cell type, cell state and post-translational modification signaling networks in organoids.
The authors present a protocol for bioengineering human intestinal mucosal grafts. This includes the isolation, expansion and biobanking of patient-derived intestinal organoids and fibroblasts and the decellularization and recellularization of human intestinal scaffolds.
The protocol describes coculture of a primary human colon monolayer derived from cells growing in colon organoids with aerotolerant bacteria, such as strains of B. thetaiotaomicron genetically engineered to respond to different stimuli in colonic microenviroments.
Skin organoids are generated from human pluripotent stem cells in a three-dimensional in vitro culture system. The mature organoids contain stratified skin layers, pigmented hair follicles, sebaceous glands, Merkel cells and sensory neurons.
The authors describe how to make both a gut-on-a-chip and a hybrid chip with a Transwell insert, and how to induce 3D morphogenesis of human intestinal epithelium from either Caco-2 cells or organoids using basolateral medium flow in both platforms.
In this Protocol Extension, Lancaster et al. describe a modified version of their original protocol (published in 2014) that can be used to reliably generate cerebral organoids of a telencephalic identity and maintain long-term viability for later stages of neural development, including axon outgrowth and neuronal maturation.
An approach to studying epithelia derived from organoids of either fetal or adult state and from both the small intestine and colon via transplantation into animals in which the colon has been damaged following administration of dextran sulfate sodium.
Takebe et al. describe a protocol for the continuous patterning of hepatic, biliary and pancreatic structures from a 3D culture of human pluripotent stem cells.
This protocol describes how to integrate organoids into a vascularized organ-on-a-chip biofabricated platform. The platform is assembled in a 96-well format and allows the connection of two tissues through a single endothelialized microchannel blood vessel.
This protocol describes the long-term culture of liver and pancreas 3D organoids from human and mouse, and differentiation of liver organoids in vitro and in vivo. Methodology for genetic manipulation of these self-renewing organoids is also detailed.
Bioengineered biliary tissue suitable for biliary reconstruction is obtained from cholangiocyte organoids derived from human primary extra- or intrahepatic duct cholangiocytes.
This protocol describes stepwise differentiation of human pluripotent stem cells into 3D kidney organoids that contain segmented nephrons connected to collecting ducts, which are surrounded by renal interstitial cells and an endothelial network.
Self-organizing 3D human blood vessel organoids are generated by mesoderm induction of hPSC aggregates and subsequent differentiation into endothelial networks and pericytes in a 3D collagen I–Matrigel matrix.
Human pluripotent stem cells are differentiated into ventral–anterior foregut spheroids and then to lung organoids, resembling the bronchi and surrounding mesenchyme of the developing human airway, or bud tip progenitor organoids.
This protocol describes a strategy for generating 3D prostate organoid cultures from healthy mouse and human prostate cells (either bulk or FAC-sorted single luminal and basal cells), metastatic prostate cancer lesions and circulating tumor cells.
This protocol describes how to isolate trophoblast from first-trimester human placentas and grow it long term in a three-dimensional organoid culture system.
Human pluripotent stem cell aggregates are formed, embedded in Matrigel and directed to differentiate to heart-forming organoids by the chemical WNT pathway modulators CHIR99021 and IWP2.