Pluripotent Stem Cell and Organoid Core
The objective of this core is to facilitate and support the development of state-of-the-art human pluripotent stem cell (hPSC) and organoid technologies by CuSTOM investigators. The services are provided by the staff of the Cincinnati Children’s Pluripotent Stem Cell Facility (PSCF) led by Dr. Chris Mayhew and Dr. James Wells, and Transgenic Animal and Genome Editing Core (TAGE) led by Dr. Yueh-Chiang Hu.
- Access to quality-controlled hPSCs and reagents
- Generation of human induced pluripotent stem cells (iPSCs)
- Consenting and generation of patient derived iPSCs
- CRISPR/Cas9-mediated genome editing of hPSCs
- Directed differentiation of hPSCs to human intestinal organoids (HIOs) and human liver organoids (HLO)
- Hands-on training courses in hPSC culture methods, iPSC generation, and HIO generation
iPSC generation for CuSTOM members
iPSCs are pluripotent cells that can be generated from any individual using techniques that are now routine in the PSCF. iPSCs have the intrinsic potential to form any human cell type and represent an important resource for basic and translational research. Furthermore, the development of methods to differentiate iPSCs to 3D organoids permits the formation of complex patient-specific tissues, facilitating development of novel models to study human development, homeostasis and disease in a highly physiologically-relevant context.
- The PSCF offers iPSC generation from fibroblasts or peripheral blood mononuclear cells (PBMCs) on a fee-for-service basis.
- CuSTOM investigators will receive subsidized rates for iPSC generation. Please contact us for current pricing.
- Investigators are responsible for acquiring the patient sample and delivering freshly isolated blood or low-passage fibroblast cultures to the PSCF for reprogramming.
- The reprogramming process typically takes 4-5 months to compete and generates 3-6 discrete clones expanded and cryopreserved at ~ p10 (6 vials of each).
- Assays for iPSC authentication and quality assessment (karyotyping and/or teratoma formation) are available for additional cost.
Patient derived iPSCs
- CuSTOM investigators have access to a streamlined, IRB-approved consenting process for acquisition of donor patient tissue samples.
- The Pluripotent Stem Cell and Organoid Core will generate de-identified patient-derived iPSC for disease modeling and preclinical studies.
- CuSTOM investigators have access to GMP grade iPSC generation.
- CuSTOM investigators will have access to a bank of iPSCs representing variety of disease states
For more information of patient derived iPSC generation and pricing contact.the core at PSCF@cchmc.org.
Genome editing iPSCs for CuSTOM members
The core offers state-of-the-art CRISPR/Cas9 genome editing of human iPSCs including indel mutations, knockin of exogenous sequences and precision editing. For example, CRISPR/Cas9-mediated correction of a single point mutation in an iPSC line derived from an individual with a genetic disease can be used to generate an isogenic pair of iPSCs differing only in the disease-associated mutation. Conversely, iPSCs harboring a disease-relevant mutation can be generated by introducing the mutation to “control” iPSCs derived from a healthy donor. Finally, the efficiency of introducing larger genetic elements such as reporter constructs is significantly enhanced by CRISPR/Cas9.
In collaboration with the Cincinnati Children's Transgenic Animal and Genome Editing (TAGE) core, the PSCF offers a full-service CRISPR/Cas9-mediated iPSC genome editing on a fee-for-service basis.
- The genome editing service consists of:
o CRISPR/Cas9 reagent design, synthesis and validation
o Transfection of iPSCs and generation of gene edited iPSC clones
o Clone genotyping
o Clone expansion and cryopreservation
- CuSTOM investigators will receive subsidized rates for iPSC genome editing.
- A typical knockout (INDEL induction) or ssODN-mediated knockin genome editing project takes 3-4 months. Projects involving generation of targeting plasmids for introduction of larger DNA fragments such as fluorescent reporters can take up to 6 months.
Please contact us for current pricing and to discuss your specific project.
PSCF subsidies for CuSTOM members
CuSTOM members receive subsidized rates for iPSC generation and genome editing services at the Pluripotent Stem Cell and Organoid Core. Please contact us for current pricing.
Christopher Mayhew, Ph.D.
Location of the Core: R3023
Remember to acknowledge the use of the core facilities in all publications. Examples of appropriate language include: “This work was supported by the Pluripotent Stem Cell and Organoid Core in the Center for Stem Cell and Organoid Medicine (CuSTOM) at Cincinnati Children’s Hospital”. If the genome editing service is received, please also acknowledge the Transgenic Animal and Genome Editing Core for the design and preparation of the CRISPR/Cas9 reagents and the genotyping service.