Organoid and 3D Cell Culture Reagents

Protocols to generate 3D brain organoids from ESCs and iPSCs were first published in 2009. These studies showed that pluripotent stem cells could differentiate into cerebral organoids containing specific cortical regions, neural progenitor populations, and cortical layer patterning. Cerebral organoids have since been employed to uncover evolutionary differences in brain development between species, mechanisms of brain region interconnectivity, and the developmental physiology of normal and diseased brain regions. iPSC-derived organoids show great potential for use in drug discovery as well as modeling neurodegenerative disease and viral brain infection.

Common Reagents for Brain Organoid Culture

Recommended Markers for Brain Organoids

Notable Published Protocols

Cancer organoids should be cultured in similar conditions to those for which the parent organoid can be grown. In some instances mutations occur in cancer stem cells that allow them to grow in the absence of normal growth factors, such as EGF or FGF. Depending on the cancer organoid it may be possible to grow them in a medium where one or more factors are removed from the normal organoid growth medium.

Notable Published Protocols

The stomach contains Lgr5+ adult stem cells that can be isolated, cultured, and differentiated in vitro into gastric organoids. Early organoid models elucidated molecular mechanisms underlying gastric development, including signaling pathways that influence fundic or antral gastric epithelium formation. Gastric organoid cultures are powerful models to study normal and diseased gastric physiology as well as more complex life science models for drug discovery and disease modeling.

Common Reagents for Gastric Organoid Culture

Recommended Markers for Gastric Organoids

Notable Published Protocols

In vitro generation of cardiac tissue is enabling advancements in drug discovery and toxicity testing, as well as facilitating the engineering of cardiac tissue for regenerative therapies. Various methods have been employed to generate 3D cardiac tissue, including iPSC-derived cardiomyocyte spheroids and bioprinting of cardiac organoids from iPSCs with subsequent differentiation into cardiomyocytes. However, protocol and reagent advancements are still needed to enhance the maturity and complexity of the cardiac tissue.

Common Reagents for Cardiac Organoid Culture

Recommended Markers for Cardiac Organoids

Notable Published Protocols

Pluripotent stem cell-derived inner ear organoids are rapidly advancing our understanding of inner ear development and physiology. Inner ear organoids have been shown to develop sensory epithelium containing the necessary hair cells, supporting cells, and synaptic-like structures that support auditory or gravitational transduction. These models have great potential for translational research, uncovering molecular and cellular mechanisms that support the regeneration of cochlear and vestibular sensory tissue.

Common Reagents for Inner Ear Organoid Culture

Notable Published Protocols

The small intestine, large intestine and colon consist of a multicellular epithelium with distinct morphological structures, including villi and invaginated crypt structures. Intestinal crypts house Lgr5+ intestinal adult stem cells that are responsible for the continuous renewal of intestinal epithelium. Intestinal crypts were first utilized to create long-term 3D culture models of the intestine, termed human intestinal organoids (hIO) or epithelial organoids. Intestinal organoid cultures are employed to study normal and diseased physiology, including barrier functions, nutrient uptake, and tissue renewal. In addition, human intestinal organoids can be generated from iPSCs. iPSC-derived hIOs have been utilized as advanced models for gastrointestinal developmental biology, drug toxicity, and personalized medicine applications.

Common Reagents for Intestinal Organoid Culture

Recommended Markers for Small Intestinal Epithelial Organoids

Recommended Markers for Colon Epithelial Organoids

Notable Published Protocols

Using pluripotent stem cells, kidney organoid culturing protocols have shown the ability to recapitulate the organ’s complex tissue cytoarchitecture, including expression of cellular markers for podocytes, proximal tubules, and distal tubules. Success in cultivating kidney organoids has facilitated research interrogating kidney development, physiology, and mechanisms underlying kidney disease (i.e. chronic kidney disease). In addition, kidney organoid research has demonstrated its potential as a translational method for kidney tissue regeneration.

Common Reagents for Kidney Organoid Culture

Recommended Markers for Kidney Organoids

Notable Published Protocols

The liver is the primary organ system for drug metabolism and detoxification. In this role, it is also highly susceptible to damage from pharmaceuticals and other chemical toxicants. Animal models and traditional in vitro assays modeling liver metabolism often fail to recapitulate the in vivo toxicity of drugs in human patients. Liver organoids, derived from primary tissue or induced pluripotent stem cells, have emerged as more complex and predictive models for hepatotoxicity and drug screening.

Common Reagents for Liver Organoid Culture

Recommended Markers for Liver Organoids

Notable Published Protocols

3D cell culture models of the pulmonary system are increasingly utilized to study lung regeneration, model disease (i.e. cystic fibrosis), and investigate mechanisms of viral lung infection (i.e. SARS-CoV-2). While lung organoids were first generated using Lgr5+ stem cells isolated from primary tissue, protocols for culturing iPSC-derived lung organoids have increased the flexibility and accessibility of this model system for use in personalized medicine and drug discovery.

Common Reagents for Lung Organoid Culture

Recommended Markers for Lung Organoids

Notable Published Protocols

Protocols to generate mammary organoids from primary epithelial tissues are helping elucidate the cell fate decisions and molecular mechanisms of mammary gland development, including ductal formation and transformation of milk-producing alveoli. Most importantly, these 3D culture techniques have enabled the cultivation of breast organoids, which are being employed for in vitro drug discovery and personalized drug screening for breast cancer.

Notable Published Protocols

Pancreatic organoids have become an informative in vitro model to study pancreatic cancer, exocrine disease, and the basic development of pancreatic ductal epithelium for potential use as regenerative or therapeutic treatment of diabetes. While robust protocols for pancreatic organoid generation using mouse primary pancreatic ductal tissues exist, protocols that support the long-term cultivation of pancreatic organoids from human tissues are still emerging.

Common Reagents for Pancreatic Organoid Culture

Recommended Markers for Pancreatic Organoids

Notable Published Protocols

Prostate organoids have rapidly evolved our ability to investigate prostate cancer, its underlying mechanisms, and novel therapeutic strategies for treatment. Prostate organoids, derived from dissociated tissues, adult stem cells, or pluripotent stem cells, are characterized as spheroid structures with differentiated, pseudostratified epithelial cell layers and express functional androgen receptors. Patient‐derived organoids, such as xenograft (PDX)‐prostate organoids, provide useful model for drug discovery and toxicology screening of potential therapeutics. Learn more about prostate cancer screening tools.

Common Reagents for Prostate Organoid Culture

Recommended Markers for Prostate Organoids

Notable Published Protocols

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Miller, A.J., Dye, B.R., Ferrer-Torres, D. et al. Generation of lung organoids from human pluripotent stem cells in vitro. Nat Protoc 14, 518–540 (2019).

Puschhof, J., Pleguezuelos-Manzano, C., Martinez-Silgado, A. et al. Intestinal organoid cocultures with microbes. Nat Protoc 16, 4633–4649 (2021).

Sander, V. Przepiorski, A. et al. Protocol for Large-Scale Production of Kidney Organoids from Human Pluripotent Stem Cells. STAR Protoc. 1(13): 100150 (2020).

Volmert, B., Kiselev, A., Juhong, A. et al. A patterned human primitive heart organoid model generated by pluripotent stem cell self-organization. Nat Commun 14, 8245 (2023).