Using Intestinal Organoids for Intestinal and Stem Cell Research


Intestinal organoids provide a major advancement in the way we study intestinal and adult stem cell biology. When intestinal crypts containing LGR5+ intestinal stem cells (ISCs) are placed in a specialized medium that simulates the intestinal stem cell niche, the ISCs rapidly proliferate and differentiate into all of the major cell types found in the intestinal epithelium: Paneth cells, goblet cells, enteroendocrine cells, and enterocytes. Remarkably, these cells arrange themselves into a crypt-villus structure surrounding a functional central lumen, thus maintaining the stem cell pool while mimicking the physiology of the adult intestinal epithelium. The resulting organoids (or “mini-guts”) can be used for a variety of research applications including:

●    intestinal stem cell biology
●    intestinal disease
●    adult stem cell biology
●    gene therapy
●    preclinical drug screening
●    personalized medicine

As an in vitro system that is capable of providing in vivo insight, intestinal organoids signify an exciting new chapter for research in these fields. More details about this culture system and its research applications can be found in our previous post, Intestinal Organoid Culture: A Convenient and Physiologically Relevant Model for Intestinal and Stem Cell Research.

Have any questions about incorporating intestinal organoids into your research?

During this Ask the Expert session, Ryan Conder, Senior Scientist at STEMCELL Technologies, will be answering your questions about intestinal organoid culture, its research applications, and IntestiCult™ Organoid Growth Medium (Mouse), the first-ever complete, defined and serum-free medium for growing intestinal organoids from mouse intestinal crypts.

In this video Ryan provides a brief overview of the topic:

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Question 1

Do you recommend using EB or monolayer culture in this application?

IntestiCult™ Organoid Growth Medium (Mouse) is optimized to provide an environment that supports the growth of intestinal stem cells located at the base of the intestinal crypts or dissociated into single cells. We recommend using primary crypts derived from the small intestine as the medium allows these crypts to grow into 3D multicellular structures, known … Continued

Question 2

I accidentally forgot to passage my organoid culture for over a week. Is it still good or do I have to start over?

Unlike other tissue culture, intestinal organoid culture can often survive for weeks without passaging, so you can continue using it. However, we recommend passaging every 7-10 days to ensure that the organoids remain healthy. Total: 0 0 0 0

Question 3

Is it true that intestinal organoids can be used for personalized medicine for treating intestinal disease? Is this only in the research stage or have clinical trials been done?

Intestinal crypts can be isolated from a patient, grown into organoids, and subjected to preclinical drug screening to determine the optimal treatment for that individual. Currently, The HUB Foundation for Organoid Technology is generating a library of human organoids, or “biobank”, derived from healthy and diseased tissue for the purposes of preclinical drug screening. Note … Continued

Question 4

We use our own homemade medium that is based on the recipe published by Toshiro Sato and Hans Clevers (Sato et al., 2009). What is different about the IntestiCult medium – what’s in it?

IntestiCult™ Organoid Growth Medium (Mouse) is an optimized version of the published medium (1), and has been developed in collaboration with Hans Clevers and The HUB Foundation for Organoid Technology. STEMCELL Technologies is the exclusive manufacturer and worldwide distributor of this complete, defined and serum-free medium for mouse intestinal organoids. The optimized recipe allows for … Continued

Question 5

If I understand the application correctly, it seems to be primarily in the research field, not in actual Cell Therapy use. If so, what is the advantage of serum-free over using serum in the culture if it is for research only?

As with all therapeutics, primary research is essential to ensure any procedure is safe and effective as a treatment. Experiments have already been performed demonstrating the feasibility of re-integration of intestinal cells in mice (1). This serves as a proof-of-concept for future, more advanced treatments requiring that the culture medium is serum-free and defined. The … Continued

Question 6

What vessels do you recommend for your 3D culture?

The culture vessels used can change depending on the researcher’s needs. For intestinal organoid culture we most commonly use 24-well culture plates but have also used 96-well plates for culture from single cells and 6-well plates for expanding cultures. You can find more details regarding the materials required in our Technical Bulletin. Total: 0 0 … Continued

Question 7

In terms of using this in drug discovery/testing, how difficult would it be for a team of cell culture scientists with minimal experience in working with stem cells to get this up and running?

Both the detailed procedure and IntestiCult™ Organoid Growth Medium (Mouse) have been designed with that thought in mind. The step-by-step manual is easy to follow and the robustness of the medium allows for a bit of a “learning curve”. Although mastering any new skill takes some practice, the time to get “up and running” with … Continued

Question 8

Do you think this technology has the potential to eliminate or reduce animal testing?

Yes, this technology definitely has the potential to reduce animal testing. Many experiments from developmental biology to drug screening can be performed on organoids rather than primary mouse culture. Organoids offer a “clean” system that allows for testing of drug effects on the intestinal epithelium alone, without any potential side-effects seen when doing animal testing. … Continued

Question 9

How finicky are these cells? Would you say easy or challenging to culture?

The primary culture of these cells is sometimes finicky as many variables regarding the mice affect the cells, and we have optimized IntestiCult™ Organoid Growth Medium (Mouse) to account for this. As for easy or challenging that can depend on your definition. On one hand culturing organoids is easy as you can be confident that … Continued

Question 10

Do you recommend Matrigel for matrix?

It is necessary that the intestinal stem cells are cultured in a specialized environment that includes a basement membrane extract to mimic a stem cell niche. We have performed all our testing using Matrigel and find it to be a suitable matrix when using IntestiCult™ Organoid Growth Medium (Mouse). Total: 0 0 0 0

Question 11

Do you feel antibiotics are necessary in the culture, I would prefer not to use them.

Antibiotics are only necessary as a preventative measure in the culture. The medium is not affected positively or negatively by their addition. Intestinal culture is inherently a very non sterile tissue and that is why we recommend the addition of antibiotics. But they can be cultured without antibiotics if you prefer not to use them. … Continued

Question 12

If I don’t see organoid growth after 4 days, should I start over or give the culture more time? How long should I wait before starting over?

You will almost always see growth within 4 days of standard culture. Different mouse models will introduce some variability into this time frame but one should be able to recognize growing epithelial cysts after 1-2 days. Living cysts can be identified by a monolayered epithelium when observed under a brightfield microscope compared to single cells … Continued

Question 13

Culture medium for colonic organoids differs from that for small intestinal organoids (e.g. Wnt3a growth factor). Does your medium contain these extra growth factors, or do they need to be added separately? Also, does the described isolation protocol differ between small intestinal and colon organoids?

Indeed the culture of colon crypts differs from that of the small intestine. We have found that IntestiCult™ Organoid Growth Medium (Mouse) works on both tissues. In the primary culture, the efficiency of cultured colon crypts that grow into mature organoids may not be as high as the efficiency of the small intestinal cultures, but … Continued