STEMdiff™ Cerebral Organoid Kit

Culture medium kit for establishment and maturation of human cerebral organoids
STEMdiff™ Cerebral Organoid Maturation Kit

Culture medium kit for extended maturation of human cerebral organoids

1 Kit
Catalog # 08571
112 USD
STEMdiff™ Cerebral Organoid Kit

Culture medium kit for establishment and maturation of human cerebral organoids

1 Kit
Catalog # 08570
328 USD
Required Products
  1. Gentle Cell Dissociation Reagent
    Gentle Cell Dissociation Reagent

    cGMP, enzyme-free cell dissociation reagent

  2. Y-27632 (Dihydrochloride)
    Y-27632

    RHO/ROCK pathway inhibitor; Inhibits ROCK1 and ROCK2

  3. D-PBS Without Ca++ and Mg++
    D-PBS (Without Ca++ and Mg++)

    Dulbecco’s phosphate-buffered saline without calcium and magnesium

  4. Costar® 24-Well Flat-Bottom Plate, Tissue Culture-Treated
    Costar® 24-Well Flat-Bottom Plate, Tissue Culture-Treated

    Clear polystyrene flat-bottom, tissue culture-treated multiwell cell culture plate with lid

  5. 6-Well Ultra-Low Adherent Plate|38071
    Ultra-Low Adherent Plate for Suspension Culture

    Sterile, flat-bottom plate, with ultra-low adherent surface for suspension cultures, with lid; 6-well format

Overview

Generate self-organized, pluripotent stem cell (PSC)-derived neural organoids with a cellular composition and structural organization representative of the developing human brain.

These defined, serum-free cell culture media and simple, four-stage protocol are based on the formulation published by Lancaster et al. (Lancaster MA et al. Nature, 2013 and Lancaster MA et al. Science, 2014) to more reliably generate cerebral organoids. Beginning with an embryoid body (EB) formation step followed by expansion of neuroepithelia, organoids generated using STEMdiff™ Cerebral Organoid Kit feature cortical-like regions including the ventricular zone (PAX6+/SOX2+/Ki-67+), outer subventricular zone (Ki-67+/p-Vimentin+), intermediate zone (TBR2+), and cortical plate (CTIP2+/MAP2+/TBR1+), which layer in similar orientations as those observed in vivo.

For extended culture periods (> 40 days), the components required for maturation can be purchased as STEMdiff™ Cerebral Organoid Maturation Kit (Catalog #08571).
Components
  • STEMdiff™ Cerebral Organoid Kit (Catalog #08570)
    • STEMdiff™ Cerebral Organoid Basal Medium 1,100 mL
    • STEMdiff™ Cerebral Organoid Basal Medium 2, 250 mL
    • STEMdiff™ Cerebral Organoid Supplement A, 10 mL
    • STEMdiff™ Cerebral Organoid Supplement B, 0.5 mL
    • STEMdiff™ Cerebral Organoid Supplement C, 0.25 mL
    • STEMdiff™ Cerebral Organoid Supplement D, 0.5 mL
    • STEMdiff™ Cerebral Organoid Supplement E, 4.5 mL
  • STEMdiff™ Cerebral Organoid Maturation Kit (Catalog #08571)
    • STEMdiff™ Cerebral Organoid Basal Medium 2, 250 mL
    • STEMdiff™ Cerebral Organoid Supplement E, 4.5 mL
Subtype
Specialized Media
Cell Type
Neural Cells, PSC-Derived, Neural Stem and Progenitor Cells, Pluripotent Stem Cells
Species
Human
Application
Cell Culture, Differentiation, Organoid Culture
Brand
STEMdiff
Area of Interest
Disease Modeling, Neuroscience, Stem Cell Biology
Formulation
Serum-Free

Scientific Resources

Product Documentation

Document Type Product Name Catalog # Lot # Language
Document Type
Product Information Sheet
Product Name
STEMdiff™ Cerebral Organoid Maturation Kit
Catalog #
08571
Lot #
All
Language
English
Document Type
Product Information Sheet
Product Name
STEMdiff™ Cerebral Organoid Kit
Catalog #
08570
Lot #
All
Language
English
Document Type
Safety Data Sheet 1
Product Name
STEMdiff™ Cerebral Organoid Maturation Kit
Catalog #
08571
Lot #
All
Language
English
Document Type
Safety Data Sheet 2
Product Name
STEMdiff™ Cerebral Organoid Maturation Kit
Catalog #
08571
Lot #
All
Language
English
Document Type
Safety Data Sheet 3
Product Name
STEMdiff™ Cerebral Organoid Maturation Kit
Catalog #
08571
Lot #
All
Language
English
Document Type
Safety Data Sheet 4
Product Name
STEMdiff™ Cerebral Organoid Maturation Kit
Catalog #
08571
Lot #
All
Language
English
Document Type
Safety Data Sheet 5
Product Name
STEMdiff™ Cerebral Organoid Maturation Kit
Catalog #
08571
Lot #
All
Language
English
Document Type
Safety Data Sheet 6
Product Name
STEMdiff™ Cerebral Organoid Maturation Kit
Catalog #
08571
Lot #
All
Language
English
Document Type
Safety Data Sheet 1
Product Name
STEMdiff™ Cerebral Organoid Kit
Catalog #
08570
Lot #
All
Language
English
Document Type
Safety Data Sheet 2
Product Name
STEMdiff™ Cerebral Organoid Kit
Catalog #
08570
Lot #
All
Language
English
Document Type
Safety Data Sheet 3
Product Name
STEMdiff™ Cerebral Organoid Kit
Catalog #
08570
Lot #
All
Language
English
Document Type
Safety Data Sheet 4
Product Name
STEMdiff™ Cerebral Organoid Kit
Catalog #
08570
Lot #
All
Language
English
Document Type
Safety Data Sheet 5
Product Name
STEMdiff™ Cerebral Organoid Kit
Catalog #
08570
Lot #
All
Language
English
Document Type
Safety Data Sheet 6
Product Name
STEMdiff™ Cerebral Organoid Kit
Catalog #
08570
Lot #
All
Language
English
Document Type
Safety Data Sheet 7
Product Name
STEMdiff™ Cerebral Organoid Kit
Catalog #
08570
Lot #
All
Language
English

Educational Materials (34)

Brochure
Organoids
Brochure
2019-2020 Cell Culture Training Catalog
Brochure
hPSC-Derived Neural Cell Research
Brochure
STEMCELL Neural Product Portfolio
Brochure
STEMdiff™ Cerebral Organoid Kit
Technical Bulletin
Cryogenic Tissue Processing and Section Immunofluorescence of Cerebral Organoids
Protocol
Neural Organoid Culture
Protocol
How to Dissociate 3D Neural Organoids into a Single-Cell Suspension
Wallchart
Cell-Reprogramming Technology and Neuroscience
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
Video
How to Grow Cerebral Organoids from Human Pluripotent Stem Cells
9:18
How to Grow Cerebral Organoids from Human Pluripotent Stem Cells
Video
STEMCELL Journal Club: Cerebral Organoids for Human-Specific Infection Modeling
28:45
STEMCELL Journal Club: Cerebral Organoids for Human-Specific Infection Modeling
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
Cerebral Organoids as 3D, Stem Cell-Derived Models of Tuberous Sclerosis Complex
43:00
Cerebral Organoids as 3D, Stem Cell-Derived Models of Tuberous Sclerosis Complex
Webinar
Brains in a Dish: Using Cerebral Organoids to Study Human Brain Development and Disease
25:30
Brains in a Dish: Using Cerebral Organoids to Study Human Brain Development and Disease
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
CRISPR-Cas9 Gene Editing of Cerebral Organoids to Model Microcephaly
10:08
CRISPR-Cas9 Gene Editing of Cerebral Organoids to Model Microcephaly
Webinar
Exploring the Impact of SARS-CoV-2 Infection on the Central Nervous System
1:00:11
Exploring the Impact of SARS-CoV-2 Infection on the Central Nervous System
Webinar
Madeline Lancaster on Brain Organoids: Modeling Human Brain Development in a Dish
47:18
Madeline Lancaster on Brain Organoids: Modeling Human Brain Development in a Dish
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
ISSCR Innovation Showcase: Advanced Brain Organoid Co-Culture Systems
1:01:37
ISSCR Innovation Showcase: Advanced Brain Organoid Co-Culture Systems
Webinar
hPSC Quality: Essential Considerations for Gene Editing, Cloning, Maintenance and Disease Modeling
50:49
hPSC Quality: Essential Considerations for Gene Editing, Cloning, Maintenance and Disease Modeling
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
CRISPR-Cas9 Gene Editing Of CDK5RAP2 In Human Pluripotent Stem Cells, Derivation Of Genetically Stable Clonal Lines And Formation Of Cerebral Organoids
Scientific Poster
STEMdiff™ Cerebral Organoid Kit: A New Tool for the Culture of 3D Brain Organoids Derived from hPSCs
Scientific Poster
STEMdiff Cerebral Organoid Kit: A New Tool for the Culture of 3D Brain Organoids Derived from Human Pluripotent Stem Cells
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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.

(A) A representative phase-contrast image of a whole cerebral organoid at day 40 generated using the STEMdiff™ Cerebral Organoid Kit. Cerebral organoids at this stage are made up of phase-dark structures that may be surrounded by regions of thinner, more translucent structures that display layering (arrowheads). (B) Immunohistological analysis on cryosections of cerebral organoids reveals cortical regions within the organoid labelled by the apical progenitor marker PAX6 (red) and neuronal marker β-tubulin III (green). (C-F) Inset of boxed region from (B). (C) PAX6+ apical progenitors (red, enclosed by dotted line) are localized to a ventricular zone-like region. β-tubulin III+ neurons (green) are adjacent to the ventricular zone. (D) CTIP2, a marker of the developing cortical plate, co-localizes with β-tubulin III+ neurons in a cortical plate-like region. Organization of the layers recapitulates early corticogenesis observed during human brain development. (E) Proliferating progenitor cells labeled by Ki-67 (green) localize along the ventricle, nuclei are counterstained with DAPI (blue). (F) An additional population of Ki-67+ cells is found in an outer subventricular zone-like region (arrowheads). Scale Bar = (A) 1 mm, (B) 500 µm and (C-F) 200 µm.

Figure 2.

Principal component analysis of hPSC and cerebral organoid transcriptomes. Cerebral organoids generated using the STEMdiff™ Cerebral Organoid Kit (filled blue circles) cluster together, and cluster with previously published (C Luo et al. Cell Rep, 2016) cerebral organoids (open blue circles). The first principal component accounts for the majority of variance seen (PC1; 80%) and distinguishes the cerebral organoid samples from the hPSCs (green circles). The second principal component accounts for only 9% of the variation, and highlights the modest expression differences between cultured organoids and primary embryonic fetal brain samples (19 post-conceptional weeks, brown circles).

Figure 3.

Heatmap of expression levels for genes associated with synaptic transmission function and neurogenesis in Day 40 organoids. These data show that gene expression of cerebral organoids generated from the STEMdiff™ Cerebral Organoid Kit are similar to published results (C Luo et al. Cell Rep, 2016).

Immunocytochemistry image of a cerebral organoid cultured in mTeSR™ Plus and directed to cerebral organoids using the STEMdiff™ Cerebral Organoid Kit.

Figure 4. Generation of Cerebral Organoids from hPSCs Maintained in mTeSR™ Plus

Human ES (H9) cells were cultured with mTeSR™ Plus and directed to cerebral organoids using the STEMdiff™ Cerebral Organoid Kit. Image shows apical progenitor marker SOX2 (purple) and neuronal marker TBR1 (green).

Publications (8)

Science (New York, N.Y.) 2020 Human CNS barrier-forming organoids with cerebrospinal fluid production. L. Pellegrini et al.

Abstract

Cerebrospinal fluid (CSF) is a vital liquid, providing nutrients and signaling molecules and clearing out toxic by-products from the brain. The CSF is produced by the choroid plexus (ChP), a protective epithelial barrier that also prevents free entry of toxic molecules or drugs from the blood. Here, we establish human ChP organoids with a selective barrier and CSF-like fluid secretion in self-contained compartments. We show that this in vitro barrier exhibits the same selectivity to small molecules as the ChP in vivo and that ChP-CSF organoids can predict central nervous system (CNS) permeability of new compounds. The transcriptomic and proteomic signatures of ChP-CSF organoids reveal a high degree of similarity to the ChP in vivo. Finally, the intersection of single-cell transcriptomics and proteomic analysis uncovers key human CSF components produced by previously unidentified specialized epithelial subtypes.
Analytical chemistry 2020 One-Stop Microfluidic Assembly of Human Brain Organoids To Model Prenatal Cannabis Exposure. Z. Ao et al.

Abstract

Prenatal cannabis exposure (PCE) influences human brain development, but it is challenging to model PCE using animals and current cell culture techniques. Here, we developed a one-stop microfluidic platform to assemble and culture human cerebral organoids from human embryonic stem cells (hESC) to investigate the effect of PCE on early human brain development. By incorporating perfusable culture chambers, air-liquid interface, and one-stop protocol, this microfluidic platform can simplify the fabrication procedure and produce a large number of organoids (169 organoids per 3.5 cm × 3.5 cm device area) without fusion, as compared with conventional fabrication methods. These one-stop microfluidic assembled cerebral organoids not only recapitulate early human brain structure, biology, and electrophysiology but also have minimal size variation and hypoxia. Under on-chip exposure to the psychoactive cannabinoid, $\Delta$-9-tetrahydrocannabinol (THC), cerebral organoids exhibited reduced neuronal maturation, downregulation of cannabinoid receptor type 1 (CB1) receptors, and impaired neurite outgrowth. Moreover, transient on-chip THC treatment also decreased spontaneous firing in these organoids. This one-stop microfluidic technique enables a simple, scalable, and repeatable organoid culture method that can be used not only for human brain organoids but also for many other human organoids including liver, kidney, retina, and tumor organoids. This technology could be widely used in modeling brain and other organ development, developmental disorders, developmental pharmacology and toxicology, and drug screening.
Scientific reports 2019 nov Extracellular Membrane Vesicles from Lactobacilli Dampen IFN-$\gamma$ Responses in a Monocyte-Dependent Manner. M. Mata Forsberg et al.

Abstract

Secreted factors derived from Lactobacillus are able to dampen pro-inflammatory cytokine responses. Still, the nature of these components and the underlying mechanisms remain elusive. Here, we aimed to identify the components and the mechanism involved in the Lactobacillus-mediated modulation of immune cell activation. PBMC were stimulated in the presence of the cell free supernatants (CFS) of cultured Lactobacillus rhamnosus GG and Lactobacillus reuteri DSM 17938, followed by evaluation of cytokine responses. We show that lactobacilli-CFS effectively dampen induced IFN-$\gamma$ and IL-17A responses from T- and NK cells in a monocyte dependent manner by a soluble factor. A proteomic array analysis highlighted Lactobacillus-induced IL-1 receptor antagonist (ra) as a potential candidate responsible for the IFN-$\gamma$ dampening activity. Indeed, addition of recombinant IL-1ra to stimulated PBMC resulted in reduced IFN-$\gamma$ production. Further characterization of the lactobacilli-CFS revealed the presence of extracellular membrane vesicles with a similar immune regulatory activity to that observed with the lactobacilli-CFS. In conclusion, we have shown that lactobacilli produce extracellular MVs, which are able to dampen pro-inflammatory cytokine responses in a monocyte-dependent manner.
Cell stem cell 2019 aug Interconversion between Tumorigenic and Differentiated States in Acute Myeloid Leukemia. M. D. McKenzie et al.

Abstract

Tumors are composed of phenotypically heterogeneous cancer cells that often resemble various differentiation states of their lineage of origin. Within this hierarchy, it is thought that an immature subpopulation of tumor-propagating cancer stem cells (CSCs) differentiates into non-tumorigenic progeny, providing a rationale for therapeutic strategies that specifically eradicate CSCs or induce their differentiation. The clinical success of these approaches depends on CSC differentiation being unidirectional rather than reversible, yet this question remains unresolved even in prototypically hierarchical malignancies, such as acute myeloid leukemia (AML). Here, we show in murine and human models of AML that, upon perturbation of endogenous expression of the lineage-determining transcription factor PU.1 or withdrawal of established differentiation therapies, some mature leukemia cells can de-differentiate and reacquire clonogenic and leukemogenic properties. Our results reveal plasticity of CSC maturation in AML, highlighting the need to therapeutically eradicate cancer cells across a range of differentiation states.
Nature Neuroscience 2019 apr Cerebral organoids at the air–liquid interface generate diverse nerve tracts with functional output S. L. Giandomenico et al.

Abstract

Neural organoids have the potential to improve our understanding of human brain development and neurological disorders. However, it remains to be seen whether these tissues can model circuit formation with functional neuronal output. Here we have adapted air–liquid interface culture to cerebral organoids, leading to improved neuronal survival and axon outgrowth. The resulting thick axon tracts display various morphologies, including long-range projection within and away from the organoid, growth-cone turning, and decussation. Single-cell RNA sequencing reveals various cortical neuronal identities, and retrograde tracing demonstrates tract morphologies that match proper molecular identities. These cultures exhibit active neuronal networks, and subcortical projecting tracts can innervate mouse spinal cord explants and evoke contractions of adjacent muscle in a manner dependent on intact organoid-derived innervating tracts. Overall, these results reveal a remarkable self-organization of corticofugal and callosal tracts with a functional output, providing new opportunities to examine relevant aspects of human CNS development and disease.
eLife 2019 Mechanisms of hyperexcitability in Alzheimer's disease hiPSC-derived neurons and cerebral organoids vs isogenic controls. S. Ghatak et al.

Abstract

Human Alzheimer's disease (AD) brains and transgenic AD mouse models manifest hyperexcitability. This aberrant electrical activity is caused by synaptic dysfunction that represents the major pathophysiological correlate of cognitive decline. However, the underlying mechanism for this excessive excitability remains incompletely understood. To investigate the basis for the hyperactivity, we performed electrophysiological and immunofluorescence studies on hiPSC-derived cerebrocortical neuronal cultures and cerebral organoids bearing AD-related mutations in presenilin-1 or amyloid precursor protein vs. isogenic gene corrected controls. In the AD hiPSC-derived neurons/organoids, we found increased excitatory bursting activity, which could be explained in part by a decrease in neurite length. AD hiPSC-derived neurons also displayed increased sodium current density and increased excitatory and decreased inhibitory synaptic activity. Our findings establish hiPSC-derived AD neuronal cultures and organoids as a relevant model of early AD pathophysiology and provide mechanistic insight into the observed hyperexcitability.
View All Publications

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