Pluripotent Stem Cell Facility

Frequently Asked Questions

We offer access to a variety of pluripotent stem cell technologies and products. Please see the services section of the website or contact us for more details. The PSCF is also committed to adapting its services to accommodate new research. Please feel free to contact us with any ideas or suggestions.

How do I start pluripotent stem cell research at Cincinnati Children's or UC?

Initially you should contact the PSCF to discuss your project. Together we will review your objectives, discuss your options and establish how the PSCF can facilitate your research. We will also give you advice about how to obtain the necessary permissions to initiate your study. Learn more about how to get started.

Can the PSCF assist with Induced Pluripotent Stem Cell (iPSC) Technology?

Yes. We have successfully generated iPSCs from multiple human cells types, including numerous disease-specific iPSC lines. Additionally, we are able to use non-integrating episomal plasmids to generate vector-free iPSC lines. Please see our iPSC page for more details.

How do I acknowledge the PSCF in our manuscripts/presentations?

Investigators receiving services/cell lines from the PSCF are required to acknowledge the facility in publications and presentations arising from the work. The following are examples of an appropriate acknowledgments that could be used in publications: “This work was supported by the Cincinnati Children's Hospital Medical Center Pluripotent Stem Cell Facility” or "The iPSC lines used in this study were generated by the Cincinnati Children's Pluripotent Stem Cell Facility".

Which hESC lines are available to NIH funded investigators?

A list of cell lines approved for NIH funding can be found at the NIH hESC registry.

What hESC/iPSC cell lines are available through the PSCF?

NIH has developed new guidelines under which NIH will fund research in the area of human stem cells. These guidelines became effective on July 7, 2009. See the Table below for the list of cell lines currently available through the PSCF. Quality tested batches of iPSCs are also available.

Cell Line ID	Originator	NIH Funding Eligible?
WA09 (H9)	WiCell	Yes
WA01 (H1)	WiCell	Yes

What training is available in the PSCF?

We offer training in basic human pluripotent stem cell culture techniques, and advanced training in iPSC generation and differentiation of hPSCs to intestinal organoids. NOTE: ESCRO approval is NOT required to receive training within the PSCF.

How does the facility quality test hPSCs?

We perform routine quality assurance testing on our hESCs. Our hESCs are prepared in batches and each batch of undifferentiated cells is analyzed for pluripotency marker expression, functional pluripotency, karyotype and absence of mycoplasma contamination. Batch tested iPSC lines that have been assayed for pluripotency marker expression, functional pluripotency, karyotype and absence of mycoplasma contamination are also available.

Can the PSCF perform hESC/iPSC characterization/quality control test for my cells?

Yes. Routine hPSC QC assays can be performed on a fee-for-service basis. Please see our iPSC page for more details.

Can the PSCF help me to direct the differentiation of hESC/iPSC cultures?

Yes. One of our main goals is to generate differentiated cell types that can be tested in translational disease model systems. As such, we are very interested in using established protocols and developing new protocols to direct hESC differentiation to useful cell types. We are currently validating protocols to generate a variety of cell types. Please see our differentiation page for details on differentiation protocols for derivation of your cells of interest.

Does the PSCF use feeder-free culture?

Yes. We routinely use hESC-qualified matrigel (Becton Dickson) and mTeSR1 (Stem Cell Technologies) for culture of hESCs. We recommend using feeder-free culture if possible.

Where can I obtain mouse feeder cells?

We do not offer frozen vials of MEF feeders for sale. Although we do not recommend using feeder-dependent culture, for those investigators who wish to use this method, commercially available feeders can be purchased from external companies. We can provide technical assistance if feeder-dependent culture is necessary.

Are hPSCs amenable to CRISPR/Cas9-mediated gene editing?

Yes. A CRISPR/Cas9 gene editing service for hPSCs is available. The PSCF has successfully used CRISPR/Cas9 genome editing to generate: 1) hPSCs harboring INDELs for gene knockout studies, 2) hPSCs harboring single base changes for disease modeling, and 3) hPSCs containing fluorescent-reporter for assessing endogenous gene activity. We recommend that investigators use the Cincinnati Children's Transgenic Animal and Genome Editing Core to design, validate and synthesize CRISPR/Cas9 reagents and genotype putative modified clones.

Can hESCs be manipulated to stably express recombinant DNA?

Generation of stable hESC clones following transient plasmid transfection is possible but occurs with low efficiency. However, retroviral and lentiviral vectors have been used to stably express a transgene (cDNA) or small-hairpin (sh) RNA in hESCs (e.g. Zhou BY et al., Inducible and reversible transgene expression in human stem cells after efficient and stable gene transfer. Stem Cells 2007, 25(3):779-89 or Papapetrou et al., Derivation of genetically modified human pluripotent stem cells with integrated transgenes at unique mapped genomic sites. Nat Protoc 2011, 6(9): 1274-1289. We have had success generating hPSC lines constitutively- or inducibly-expressing shRNA that maintains effective knockdown during differentiation and have achieved infection of differentiated cells with recombinant adenovirus (see Spence et al, Nature. 2011 Feb 3;470(7332):105-9).

What consulting services does the PSCF offer?

We are here to help you in any aspect of your research using hESCs or iPSCs. Specifically, we are able to assist investigators with experimental design, including cell line selection and appropriate methodologies, as well as data analysis and interpretation. We are also able to provide assistance for investigators who are constructing manuscripts and grant applications pertaining to hESC research. Consulting fees are waived for investigators providing appropriate salary support for PSCF staff.

What is the ESCRO committee?

All research involving human embryonic stem cells at Cincinnati Children's/UC is subject to review by the Embryonic Stem Cell Research Oversight (ESCRO) committee. In 2005, the National Academies of Sciences published “Guidelines for Human Embryonic Stem Cell Research”, and recommended that institutions conducting hESC research establish an ESCRO committee to ensure that applicable regulatory requirements are met, and that hESC research is conducted in accordance with the highest possible ethical standards. ESCRO committees are intended to provide local oversight of all issues related to derivation and research use of hESC lines, and to facilitate education of investigators involved in hESC research. Contact the Cincinnati Children's ESCRO committee at ESCRO@cchmc.org.

When do I need to apply for ESCRO approval?

The PSCF has ESCRO approval for the majority of common hESC procedures. During preliminary discussions, the facility director will review an investigator's planned experiments and ascertain whether additional ESCRO approval is required. If not, we encourage investigators to perform their approved experiments in the facility using our ESCRO approval. This is ideal for performing proof of principle studies. However, if an investigator proposes experimental approaches not covered in our approvals, or wishes to use cells not supplied by the facility, he/she must obtain their own ESCRO approval, as well as appropriate IBC and IACUC permissions before beginning hESC research. Regardless of the level of ESCRO approval required the PSCF will assist to facilitate the application procedure. Contact the Cincinnati Children's ESCRO committee at ESCRO@cchmc.org.

How do I obtain approval to use human Embryonic Stem Cells from the PSCF?

Specific permissions must be obtained from the originating institutions to use the hESCs provided by the PSCF. Detailed descriptions of these procedures can be found here.

Can the PSCF provide services to non Cincinnati Children's / UC investigators?

Yes. Most services are available to both internal and external investigators. Please contact us for details.

Pluripotent Stem Cell Facility

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