STEMdiff™ Intestinal Organoid Kit

Serum-free medium kit for establishment and maintenance of human intestinal organoids from hPSCs
STEMdiff™ Intestinal Organoid Growth Medium

Serum-free medium kit for maintenance of hPSC-derived human intestinal organoids

1 Kit
Catalog # 05145
509 USD
STEMdiff™ Intestinal Organoid Kit

Serum-free medium system for establishment and maintenance of human intestinal organoids from hPSCs

1 Kit
Catalog # 05140
1,021 USD
Required Products
  1. mTeSR™1
    mTeSR™1

    cGMP, feeder-free maintenance medium for human ES and iPS cells

  2. Gentle Cell Dissociation Reagent
    Gentle Cell Dissociation Reagent

    cGMP, enzyme-free cell dissociation reagent

  3. AggreWell™ Rinsing Solution
    Anti-Adherence Rinsing Solution

    Rinsing solution for cultureware to prevent cell adhesion

  4. Y-27632 (Dihydrochloride)
    Y-27632

    RHO/ROCK pathway inhibitor; Inhibits ROCK1 and ROCK2

Overview

STEMdiff™ Intestinal Organoid Kit (Catalog #05140) is a serum-free cell culture medium system that enables robust and efficient generation of human pluripotent stem cell (hPSC)-derived intestinal organoids in a simple, three-stage protocol. Using this kit, hPSCs are directed through induction of definitive endoderm, mid-/hindgut spheroids, and generation of intestinal organoids that can be maintained long-term through passaging or cryopreserved. The organoids exhibit a cellular composition and organization that models the developing intestinal epithelium and associated mesenchyme, making them a convenient model system with direct relevance to the developing intestine. STEMdiff™ Intestinal Organoid Kit is based on the formulation published by Spence et al. (Nature 2011) and has been optimized to increase efficiency and reproducibility of organoid formation and expansion across hPSC lines. Human intestinal organoids can be used as a model system for studying intestinal development and cell biology, intestinal inflammation, intestinal regeneration, microbial interaction, disease modeling, drug discovery, and compound screening. The kit is optimized for differentiation of cells maintained in mTeSR™1.

For extended culture and passaging, the kit components required for organoid maintenance can be purchased as STEMdiff™ Intestinal Organoid Growth Medium (Catalog #05145).
Advantages
• Relevant model system for the developing small intestinal epithelium and associated mesenchyme
• Robust medium supports efficient differentiation of human ES and iPS cell lines to intestinal organoids
• Convenient model system that can be maintained long-term through passaging or cryopreserved for experimental flexibility
• Serum-free medium system optimized for low experimental variability
Components
  • STEMdiff™ Intestinal Organoid Kit (Catalog #05140)
    • STEMdiff™ Endoderm Basal Medium,100 mL
    • STEMdiff™ Definitive Endoderm Supplement CJ, 1.1 mL
    • STEMdiff™ Gastrointestinal Supplement PK, 0.64 mL
    • STEMdiff™ Gastrointestinal Supplement UB, 0.64 mL
    • STEMdiff™ Intestinal Organoid Basal Medium, 100 mL
    • STEMdiff™ Intestinal Organoid Supplement, 2 mL
  • STEMdiff™ Intestinal Organoid Growth Medium (Catalog #05145)
    • STEMdiff™ Intestinal Organoid Basal Medium, 100 mL
    • STEMdiff™ Intestinal Organoid Supplement, 2 mL
Cell Type
Endoderm, PSC-Derived, Intestinal Cells
Species
Human
Application
Cell Culture, Differentiation, Organoid Culture
Brand
STEMdiff
Area of Interest
Disease Modeling, Drug Discovery and Toxicity Testing, Epithelial Cell Biology
Formulation
Serum-Free

Scientific Resources

Product Documentation

Document Type Product Name Catalog # Lot # Language
Document Type
Product Information Sheet
Product Name
STEMdiff™ Intestinal Organoid Growth Medium
Catalog #
05145
Lot #
All
Language
English
Document Type
Product Information Sheet
Product Name
STEMdiff™ Intestinal Organoid Kit
Catalog #
05140
Lot #
All
Language
English
Document Type
Manual
Product Name
STEMdiff™ Intestinal Organoid Kit
Catalog #
05140
Lot #
All
Language
English
Document Type
Safety Data Sheet 1
Product Name
STEMdiff™ Intestinal Organoid Growth Medium
Catalog #
05145
Lot #
All
Language
English
Document Type
Safety Data Sheet 2
Product Name
STEMdiff™ Intestinal Organoid Growth Medium
Catalog #
05145
Lot #
All
Language
English
Document Type
Safety Data Sheet 3
Product Name
STEMdiff™ Intestinal Organoid Growth Medium
Catalog #
05145
Lot #
All
Language
English
Document Type
Safety Data Sheet 4
Product Name
STEMdiff™ Intestinal Organoid Growth Medium
Catalog #
05145
Lot #
All
Language
English
Document Type
Safety Data Sheet 5
Product Name
STEMdiff™ Intestinal Organoid Growth Medium
Catalog #
05145
Lot #
All
Language
English
Document Type
Safety Data Sheet 6
Product Name
STEMdiff™ Intestinal Organoid Growth Medium
Catalog #
05145
Lot #
All
Language
English
Document Type
Safety Data Sheet 1
Product Name
STEMdiff™ Intestinal Organoid Kit
Catalog #
05140
Lot #
All
Language
English
Document Type
Safety Data Sheet 2
Product Name
STEMdiff™ Intestinal Organoid Kit
Catalog #
05140
Lot #
All
Language
English
Document Type
Safety Data Sheet 3
Product Name
STEMdiff™ Intestinal Organoid Kit
Catalog #
05140
Lot #
All
Language
English
Document Type
Safety Data Sheet 4
Product Name
STEMdiff™ Intestinal Organoid Kit
Catalog #
05140
Lot #
All
Language
English
Document Type
Safety Data Sheet 5
Product Name
STEMdiff™ Intestinal Organoid Kit
Catalog #
05140
Lot #
All
Language
English
Document Type
Safety Data Sheet 6
Product Name
STEMdiff™ Intestinal Organoid Kit
Catalog #
05140
Lot #
All
Language
English

Educational Materials (31)

Brochure
Organoids
Brochure
Intestinal Organoids
Brochure
STEMdiff™ Intestinal Organoid Kit
Wallchart
SnapShot: Growing Organoids from Stem Cells
Wallchart
Derivation and Applications of Human Pluripotent Stem Cells
Video
How to Count Intestinal Crypts for Mouse Organoid Cultures
1:09
How to Count Intestinal Crypts for Mouse Organoid Cultures
Video
Supporting Science Globally: A look into how scientists in Singapore are using organoids
4:46
Supporting Science Globally: A look into how scientists in Singapore are using organoids
Webinar
GI Tract Organoids: Using Advanced Tissue Models to Interrogate Absorption and Regulation
24:56
GI Tract Organoids: Using Advanced Tissue Models to Interrogate Absorption and Regulation
Webinar
Organoid Expert Panel
52:35
Organoid Expert Panel
Webinar
HUB & STEMCELL Organoids as Models of Infectious Disease Mini-Symposium
1:10:09
HUB & STEMCELL Organoids as Models of Infectious Disease Mini-Symposium
Webinar
From Stem Cells to Organoids: Modeling the Gastrointestinal Tract
1:03:40
From Stem Cells to Organoids: Modeling the Gastrointestinal Tract
Webinar
Nature Research Round Table: Modeling Congenital Pediatric Diarrhea in Intestinal Enteroids
15:15
Nature Research Round Table: Modeling Congenital Pediatric Diarrhea in Intestinal Enteroids
Webinar
Nature Research Round Table: Self-Organization and Symmetry Breaking in Intestinal Organoid Development
11:52
Nature Research Round Table: Self-Organization and Symmetry Breaking in Intestinal Organoid Development
Webinar
Nature Research Round Table: Organoids as Models for Human Disease
15:10
Nature Research Round Table: Organoids as Models for Human Disease
Webinar
Spotlight on Organoids: Expert Panel
Webinar
Modeling Human Gastrointestinal Development and Disease Using Pluripotent Stem Cells
56:31
Modeling Human Gastrointestinal Development and Disease Using Pluripotent Stem Cells
Webinar
Nature Research Round Table: The Promise of Organoid Medicine
17:17
Nature Research Round Table: The Promise of Organoid Medicine
Webinar
Nature Research Round Table: Liver Organoids for the Study of Liver Regeneration and Disease
12:57
Nature Research Round Table: Liver Organoids for the Study of Liver Regeneration and Disease
Webinar
Nature Research Round Table: Modeling Intestinal Development In Vivo and In Vitro
10:17
Nature Research Round Table: Modeling Intestinal Development In Vivo and In Vitro
Webinar
Challenges in Translating iPSC Technology
41:47
Challenges in Translating iPSC Technology
Webinar
Patient-Derived Organoids for Drug Screening and Development
50:25
Patient-Derived Organoids for Drug Screening and Development
Webinar
Nature Research Round Table: Organoids as an Enabling Technology for Precision Cancer Medicine
17:20
Nature Research Round Table: Organoids as an Enabling Technology for Precision Cancer Medicine
Webinar
Nature Research Round Table: Organoid Modeling of Stem Cells and Disease Microenvironments
4:47
Nature Research Round Table: Organoid Modeling of Stem Cells and Disease Microenvironments
Webinar
Nature Research Round Table: Progress and Challenges in Organoid Models of Human Brain Development
9:36
Nature Research Round Table: Progress and Challenges in Organoid Models of Human Brain Development
Scientific Poster
Efficient Establishment and Long-term Maintenance of 3-dimensional Human Small Intestinal and Colonic Organoids Using a Novel IntestiCult™ Organoid Growth Medium (Human)
Scientific Poster
Highly Efficient Differentiation of Human Pluripotent Stem Cells into Long-term Expandable “Mini-gut” Organoids
Interview
Heather McCauley, PhD
Interview
Tamara Zietek, PhD
Interview
Caroline Lindemans, MD, PhD
Interview
Maartje van der Heijden
Interview
Jason Spence
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Product Applications

This product is designed for use in the following research area(s) as part of the highlighted workflow stage(s). Explore these workflows to learn more about the other products we offer to support each research area.

Data and Publications

Data

Figure 1. STEMdiff™ Intestinal Organoid Kit Enables the Growth of Intestinal Organoids from PSCs

STEMdiff™ Intestinal Organoid Kit facilitates directed differentiation of PSCs to form human small intestinal organoids. The organoids exhibit a polarized epithelial monolayer surrounding a hollow lumen and are associated with a mesenchymal cell population. Pictured are passage 3 human intestinal organoids generated using this kit.

Figure 2. Generation of Human Intestinal Organoid Cultures Using the STEMdiff™ Intestinal Organoid Kit

(A) Human PSC cultures progress through a three-stage differentiation process to generate human intestinal organoids. By day 3 of the protocol, cultures exhibit characteristics typical of definitive endoderm and mid-/hindgut differentiation is initiated. During mid-/hindgut differentiation (days 5 - 9), cells form mid-/hindgut spheroids that are released from the cell monolayer into the culture medium. These spheroids are collected and embedded in extracellular matrix. (B) Embedded mid-/hindgut spheroids cultured in STEMdiff™ Intestinal Organoid Growth Medium mature into intestinal organoids (days in parentheses indicate days post-embedding in a given passage). Once established, intestinal organoids can be maintained and expanded in culture by passaging every 7 - 10 days. After multiple passages, the organoids generally exhibit less sinking within the matrix dome and a lower proportion of mesenchymal cells.

Figure 3. STEMdiff™ Intestinal Organoid Kit Supports Robust Differentiation and Expansion Across ESC and iPSC Lines

STEMdiff™ Intestinal Organoid Kit enables high-efficiency generation of intestinal organoids from both ESCs (H9, H7) and iPSCs (WLS-1C, STiPS-MOO1). (A) Organoids initiated from a variety of cell lines show efficient induction of definitive endoderm, measured by co-expression of FOXA2 and SOX17 on day 3 of differentiation. (B) Both ESCand iPSC-derived cultures demonstrate efficient spheroid formation upon mid-/hindgut induction. The total number of spheroids obtained per well in a given differentiation is shown. (C) Organoids cultured from either ESCs or iPSCs can be expanded and maintained over multiple passages. Shown is the total cell yield per passage. Organoids were passaged every 7 - 10 days with a split ratio between 1 in 2 and 1 in 4. Data points represent the mean of 3 biological replicates. Error bars throughout represent standard deviation of the mean.

Figure 4. Characteristics of Mid-/Hindgut Spheroids Generated with STEMdiff™ Intestinal Organoid Kit

(A) Cultures differentiated using STEMdiff™ Intestinal Organoid Kit exhibit the expected markers during definitive endoderm and mid-/hindgut specification. During the protocol, gene expression patterns shift from pluripotency markers (day 0) to definitive endoderm markers by day 3 and those of the mid-/hindgut epithelium by day 9. Mid-/hindgut cultures (day 9) also express markers of the associated mesenchyme. Marker levels were assessed by RT-qPCR and normalized to expression levels for undifferentiatied H9 cells. (B) Mid-/hindgut spheroids (day 9) express markers of the intestinal epithelium (CDX2, E-cadherin, EPCAM). (C) Mid-/hindgut spheroids (day 9) also incorporate components of the associated mesenchyme (vimentin).

Figure 5. Intestinal Organoids Cultured In STEMdiff™ Intestinal Organoid Growth Medium Exhibit Features of the Intestinal Epithelium

(A) Differentiated PSC-derived intestinal organoids express markers of the intestinal epithelium and the associated mesenchyme. Marker levels were assessed by RT-qPCR and normalized to expression levels for undifferentiated H9 cells. (B,C) Intestinal organoids express markers of intestinal progenitor cells including CDX2 and the intestinal crypt marker SOX9. The organoids are composed of a polarized epithelium, visualized by the localization of EPCAM to the exterior (basolateral) surface of the organoids (B), and express markers typical of mature cell types including MUC2 (B: goblet cells) and CHGA (C: enteroendocrine cells). (D,E) Observation of desmin (D) and vimentin (E) in intestinal organoids demonstrates incorporation of mesenchymal cells in the organoid cultures, while KRT20 (D) and Ki67 (E) are markers of differentiated intestinal cells and putative intestinal stem cells, respectively. Images are digital cross-sections of whole-mount immunofluorescence-stained intestinal organoids at P28 (Day 7).

Immunocytochemistry image of an intestinal organoid cultured in mTeSR™ Plus and directed to intestinal organoids using the STEMdiff™ Intestinal Organoid Kit.

Figure 6. Generation of Intestinal Organoids from hPSCs Maintained in mTeSR™ Plus

Human ES (H9) cells were cultured with mTeSR™ Plus and directed to intestinal organoids using the STEMdiff™ Intestinal Organoid Kit. Image shows markers of the intestinal epithelium EpCAM (green) and CDX2 (red), and intestinal mesenchyme marker vimentin (white). Nuclei are counterstained with DAPI (blue).

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