Description

Generating human cerebral organoids from induced pluripotent stem cells can be affected by uneven embryoid body formation, variable neural induction, inconsistent extracellular matrix embedding, and insufficient nutrient exchange during long-term three-dimensional culture. These variables may influence organoid size, optical density, neural tissue expansion, and the formation of cortical-like regions.

XR Cerebral Organoid Differentiation Kit is a chemically defined, four-stage culture system designed to differentiate human pluripotent stem cells, including human induced pluripotent stem cells and human embryonic stem cells, into three-dimensional cerebral organoids. The staged workflow guides cultures through embryoid body formation, neural induction, neural tissue expansion, matrix-supported growth, and orbital maturation.

The resulting hPSC-derived cerebral organoids can develop into large, optically dense structures containing multiple neural lineages and observable cortical-like regions. This human brain organoid culture system provides researchers with a structured workflow for studying early human cortical development and characterizing three-dimensional neural tissue formation.

Each kit contains four stage-specific cerebral organoid media and supports the generation of approximately 48 organoids when used according to the recommended workflow.

Specifications

Product Name	XR Cerebral Organoid Differentiation Kit
SKU	XR-iCereOrg-Kit
Product Type	Cerebral organoid differentiation medium kit
Starting Cells	Human pluripotent stem cells, including iPSCs and hESCs
Culture Format	Embryoid body-based three-dimensional organoid culture
Differentiation Strategy	Four-stage directed differentiation workflow
Medium A	30 mL
Medium B	30 mL
Medium C	30 mL
Medium D	160 mL
Approximate Yield	Approximately 48 cerebral organoids per kit
Initial Plate Format	96-well low-attachment U-bottom plate
Expansion Plate Format	24-well low-attachment plate
Orbital Culture Speed	65–75 rpm during the maturation stage
Recommended Culture Period	40 days or longer, depending on the required organoid size and experimental endpoint
Formulation	Chemically defined, stage-specific culture system
Grade	Research use only

Kit Components

Component	Volume	Workflow Stage
Cerebral Organoid Induction Medium A	30 mL	Embryoid body formation
Cerebral Organoid Induction Medium B	30 mL	Neural induction
Cerebral Organoid Induction Medium C	30 mL	Matrix-embedded neural expansion
Cerebral Organoid Induction Medium D	160 mL	Orbital culture and organoid maturation

Features

Four-stage cerebral organoid differentiation workflow designed for human pluripotent stem cells
Chemically defined culture system for controlled stage-specific neural differentiation
Supports embryoid body formation in low-attachment U-bottom plates
Guides cultures through neural induction, matrix embedding, neural expansion, and long-term maturation
Designed to generate large, optically dense human cerebral organoids
Supports the development of multiple neural lineages and observable cortical-like regions
Includes sufficient stage-specific media for approximately 48 cerebral organoids
Compatible with extended orbital culture at 65–75 rpm from Day 14 onward
Suitable for iPSC cerebral organoid and hESC cerebral organoid research workflows

Cerebral Organoid Differentiation Workflow

The recommended protocol begins with healthy human pluripotent stem cell cultures at approximately 80% confluency. Cells are dissociated into single cells before sequential exposure to the four stage-specific media.

Stage 1: Day 0–6 — Embryoid Body Formation

Dissociate iPSCs or other hPSCs into single cells and seed them into a 96-well low-attachment U-bottom plate using Cerebral Organoid Induction Medium A. Continue EB culture through Day 6 with medium replacement every other day.

Stage 2: Day 7–10 — Neural Induction

Transfer the developing organoids into a 24-well low-attachment plate and replace the medium with Cerebral Organoid Induction Medium B. Maintain the cultures under static conditions and replace the medium every other day.

Stage 3: Day 11–13 — Matrix Embedding and Neural Expansion

Embed each developing organoid in an extracellular matrix gel and return it to a 24-well low-attachment plate. Culture with Cerebral Organoid Induction Medium C under static conditions, replacing the medium every other day.

Stage 4: Day 14–40+ — Orbital Culture and Maturation

Place the 24-well low-attachment plate on an orbital shaker at 65–75 rpm and change to Cerebral Organoid Induction Medium D. Replace the medium every other day and continue culture until the organoids reach the size required for the planned experiment.

Important: Organoid differentiation performance can be influenced by the quality, confluency, dissociation condition, viability, and pluripotency status of the starting hPSC culture. Matrix handling and orbital culture conditions should also be kept consistent between experiments.

Human iPSC cerebral organoid differentiation timeline showing embryoid body formation neural induction matrix embedding and orbital maturation

Performance and Characterization Data

Progressive Three-Dimensional Organoid Formation

The source data show a staged morphological transition from adherent iPSC culture to compact embryoid bodies, followed by neural induction, neural tissue expansion, and formation of a large cerebral organoid. The Day 40+ morphology demonstrates the continued three-dimensional growth supported by the maturation medium and orbital culture stage.

Neural and Cortical Marker Expression

Immunofluorescence images provided with the kit show expression of neural development-associated markers, including TUJ, PAX6, CTIP2, and FOXG1. Together, these markers are used to characterize neuronal differentiation, neural progenitor populations, cortical neuronal development, and forebrain-associated identity within the generated organoids.

Marker	Common Research Association
TUJ	Neuronal differentiation and neuronal structures
PAX6	Neural progenitor and neuroectoderm-associated populations
CTIP2	Deep-layer cortical neuron-associated differentiation
FOXG1	Forebrain-associated neural development

TUJ PAX6 CTIP2 and FOXG1 immunofluorescence staining in human iPSC derived cerebral organoids

Applications

XR Cerebral Organoid Differentiation Kit is intended for research workflows requiring a three-dimensional human neural model derived from pluripotent stem cells. The four-stage system supports experimental designs focused on organoid generation, early neural differentiation, cortical-like tissue organization, and characterization of hPSC-derived brain tissue.

Human cerebral organoid generation: Differentiation of human iPSCs or hESCs into three-dimensional cerebral organoids through an EB-based workflow.
Early human brain development research: Study of cellular and structural changes associated with early neural and cerebral tissue development in vitro.
Cortical development studies: Investigation of cortical-like region formation and expression of development-associated markers such as PAX6 and CTIP2.
Forebrain development research: Generation and characterization of neural tissue expressing the forebrain-associated marker FOXG1.
Neural induction studies: Evaluation of the transition from pluripotent stem cells and embryoid bodies toward neuroectodermal and neural progenitor populations.
Neuronal differentiation research: Characterization of neuronal development and TUJ-positive structures in a three-dimensional organoid environment.
Human iPSC line comparison: Comparison of cerebral organoid formation between different human induced pluripotent stem cell lines under a common staged protocol.
Gene function research: Phenotypic characterization of genetically modified hPSC lines during cerebral organoid differentiation.
Organoid morphology analysis: Assessment of EB formation, optical density, neural tissue expansion, organoid size, and structural development over time.
Long-term neural organoid culture: Continued orbital culture beyond Day 40 for studies requiring larger or more developed cerebral organoids.
Three-dimensional neural model research: Use of hPSC-derived cerebral organoids as in vitro 3D brain models for basic neurodevelopmental research.
Neural marker characterization: Immunofluorescence analysis of markers associated with neurons, neural progenitors, cortical neurons, and forebrain identity.

Key Workflow Considerations

Starting Cell Quality

Begin differentiation with healthy, actively growing hPSC cultures at approximately 80% confluency. Cultures showing substantial spontaneous differentiation, low viability, or irregular colony morphology may produce less consistent embryoid bodies.

Single-Cell Dissociation

Consistent single-cell preparation is important for generating uniform EBs in the 96-well low-attachment U-bottom plate. Excessive mechanical stress or prolonged dissociation may reduce cell viability and affect early aggregate formation.

Extracellular Matrix Embedding

During Days 11–13, developing organoids are embedded in an extracellular matrix gel. Consistent matrix volume, handling temperature, and positioning can help reduce variation in neural tissue expansion between organoids.

Orbital Culture

From Day 14 onward, orbital culture at 65–75 rpm supports medium movement around the developing organoids. Shaker speed, plate position, working volume, and medium replacement schedule should remain consistent throughout the maturation phase.

Why Choose XR Cerebral Organoid Differentiation Kit?

Cerebral organoid generation is a multistage process in which variation during EB formation, neural induction, matrix embedding, or maturation can affect the final organoid phenotype. Researchers therefore require more than a general neural differentiation medium; they need a coordinated workflow that supports each developmental stage with the appropriate culture environment.

XR Cerebral Organoid Differentiation Kit provides four stage-specific media organized around the major steps of human cerebral organoid culture. The workflow begins with controlled embryoid body formation and progresses through neural induction, matrix-supported expansion, and long-term orbital maturation. This structure helps laboratories standardize the timing and medium transitions used during iPSC cerebral organoid differentiation.

The kit also provides sufficient medium to generate approximately 48 organoids and includes characterization data for TUJ, PAX6, CTIP2, and FOXG1 expression. It is therefore suitable for laboratories developing reproducible hPSC-derived cerebral organoid workflows for early human neural and cortical development research.

FAQ

What is the XR Cerebral Organoid Differentiation Kit used for?

XR Cerebral Organoid Differentiation Kit is used to differentiate human pluripotent stem cells, including iPSCs and hESCs, into three-dimensional cerebral organoids through a four-stage culture workflow.

What types of cells can be used to generate cerebral organoids with this kit?

The kit is designed for human pluripotent stem cells, including human induced pluripotent stem cells and human embryonic stem cells. Starting cultures should be healthy and display suitable pluripotent stem cell morphology before differentiation.

Is this an iPSC cerebral organoid differentiation kit?

Yes. The system supports iPSC cerebral organoid differentiation beginning with single-cell dissociation and EB formation, followed by neural induction, matrix embedding, neural expansion, and orbital maturation.

What is included in the cerebral organoid kit?

The kit contains 30 mL of Cerebral Organoid Induction Medium A, 30 mL of Medium B, 30 mL of Medium C, and 160 mL of Medium D. Each medium is used during a different stage of the differentiation workflow.

How many human cerebral organoids can be generated with one kit?

One kit provides sufficient stage-specific media to generate approximately 48 cerebral organoids when used according to the recommended culture workflow.

How long does cerebral organoid differentiation take?

The initial workflow extends through Day 40 or longer. EB formation occurs during Days 0–6, neural induction during Days 7–10, matrix-embedded expansion during Days 11–13, and orbital maturation begins on Day 14.

Does this cerebral organoid culture workflow use embryoid bodies?

Yes. Human pluripotent stem cells are dissociated into single cells and cultured in a 96-well low-attachment U-bottom plate to form embryoid bodies before neural induction.

Is extracellular matrix embedding required?

Yes. According to the recommended protocol, developing organoids are embedded in an extracellular matrix gel during Days 11–13 before the long-term orbital maturation stage.

What orbital shaker speed is recommended for organoid maturation?

The recommended orbital culture speed is 65–75 rpm beginning on Day 14. Laboratories should maintain consistent plate position, culture volume, and shaker conditions throughout long-term cerebral organoid culture.

What markers were used to characterize the generated cerebral organoids?

The product data include immunofluorescence images for TUJ, PAX6, CTIP2, and FOXG1. These markers are commonly associated with neuronal differentiation, neural progenitor populations, cortical neuronal development, and forebrain identity.

Can the kit be used to study cortical development?

The generated organoids can contain observable cortical-like regions, and the supplied data include PAX6 and CTIP2 staining. The system is therefore suitable for research involving early cortical-like tissue development and neural differentiation.

Is the culture medium chemically defined?

Yes. XR Cerebral Organoid Differentiation Kit is described as a chemically defined, stage-specific differentiation system for human cerebral organoid culture.

Can cerebral organoids be cultured beyond Day 40?

Yes. The protocol indicates that orbital culture with Medium D can continue beyond Day 40 until the organoids reach the size required for the planned research endpoint.

Is this product intended for clinical or diagnostic use?

No. XR Cerebral Organoid Differentiation Kit is intended for research use only and is not intended for diagnostic, therapeutic, or clinical applications.

For Research Use Only. Not for use in diagnostic or therapeutic procedures.

XR iPSC-Derived Cerebral Organoid Differentiation Kit