Human Intestinal Organoid Culture Protocol
This protocol outlines a method for culturing normal human intestinal organoids based on protocols described by C. Pleguezuelos-Manzano and Hans Clevers (1). It is intended to culture organoids from normal human intestinal tissues using Cultrex™ UltiMatrix RGF Basement Membrane Extract as a scaffold, along with recombinant proteins, small molecules, and media supplements available from Bio-Techne. This protocol not only provides you with the recipe for preparing intestinal organoid culture medium, but also notes specific tips used by our in-house cell culture scientists to simplify the culture of human intestinal organoids including:
- Considerations for starting intestinal organoid cultures from frozen stocks
- Materials needed for intestinal organoid culture and a detailed recipe for preparing intestinal organoid culture medium
- Specific instructions for successfully maintaining and passaging human intestinal organoids
- Image examples of human intestinal organoids cultured from iPSCs or adult stem cells using Cultrex UltiMatrix RGF Basement Membrane Extract and stained for specific intestinal markers
- Cryobanking and preservation of intestinal organoids for future experiments
References
- Pleguezuelos-Manzano, C. et al. (2020) Curr. Protoc. Immunol. 130: e106.
Related Resources
Establishing and optimizing the appropriate conditions for culturing different types of organoids can be technically challenging. Read this application note to learn about the importance of growth factor bioactivity and reagent consistency for achieving robust, reproducible organoid growth.
Cultrex Basement Membrane Extracts and Extracellular Matrix Proteins for 2-D and 3-D Cell Culture
Explore our full line of extracellular matrix products designed and qualified for expanding, passaging, and differentiating cultures to support reproducible and robust organoid growth. These products, along with essential reagents like Cultrex Organoid Harvesting Solution, will allow you to take your 3-D cell culture research to the next level.
Recombinant Proteins for Organoid Culture
We offer a complete catalog of recombinant proteins used for culturing different types of organoids, including even the most difficult to express and purify proteins such as R-Spondin 1, R-Spondin 3, Noggin, and Wnt-3a. Our recombinant proteins frequently display higher levels of activity than the same proteins provided by leading competitors and are rigorously tested for lot-to-lot consistency to ensure that they will support optimized, reproducible organoid growth.
Browse our selection of small molecules that are commonly used as 3-D growth matrix components or additives for supporting long-term organoid growth.
Media Supplements for Organoid Research
Improve upon the traditional B27- and NS21-like supplements with our N-2 MAX and N21-MAX media supplements for organoid cultures. N-2 MAX and N21-MAX are both serum-free, fully-defined supplements that are designed to eliminate uncontrolled variables, such as those found in serum-containing media and in some commercial serum-free supplements, and they have been shown to increase the viability and health of differentiated cell types during long-term cell culture.
A Complete Collection of Resources and Reagents for Organoid Research
Explore our complete collection of resources and reagents for organoid research including culture protocols for growing different types of organoids, product peer reviews, webinars, product highlights, and eBooks.