Frequently Asked Questions for Human Stem Cell Core (HSCC)

2. What types of human embryonic stem (hES) cells are involved?
   
   This core will start with classic hES cell lines derived from the blastocyst stage of human embryos, developed from surplus embryos from in vitro fertilization procedures. Our core intends to support research using both NIH-approved hES cells (derivation occurred prior to August 9, 2001) and subsequently derived hES cell lines. It is often that hES cells are also abbreviated as hESCs.
   
   
3. I am a scientist funded by the NIH. How many cell lines are available to me, and how can I get them?
   
   As of March 2007, there are 21 independent, fully developed stem cell lines available for widespread distribution to researchers. Some of these cell lines are available through the NIH-funded National Stem Cell Bank at reduced cost ($500 per vial). The remaining lines may be purchased by contacting the cell line providers directly. The National Stem Cell Bank is currently operated by WiCell Research Institute (Madison, WI). Information on the lines and how to contact the National Stem Cell Bank and the individual providers can be found on the NIH Stem Cell Registry.
   
   In addition, this site provides researchers with a unique NIH identifier code to apply for federal funds to do research using human embryonic stem cells.
   
   US Federal funding can be only used to study NIH-approved hES cell lines. However, Maryland and private funding could be used for research involving either NIH or non-NIH hES cell lines, provided permission is obtained from institutional regulatory bodies.
   
   
4. How do I start hES cell research at Johns Hopkins University?
   
   First, you need to obtain an approval from Embryonic Stem Cell Research Oversight Committee (ESCRO) at the JHU SOM.
   
   Second, you need to finish material transfer agreement to obtain hES cells from National Stem Cell Bank or other hES cell providers.
   
   
5. How do I begin the MTA process at JHU?
   
   Please contact:
   Julia M. Brill, Esq.
   Portfolio Director
   Johns Hopkins Technology Transfer
   100 North Charles Street, Suite 500
   Baltimore, MD 21201
   
   Julia Brill - [email protected]
   Tel 410.516.4971
   Fax 410.516.6499
   
   
6. I'm interested in purchasing more than one cell line from the NIH Stem Cell Registry. What is known about the status of the cell lines and their availability?
   
   Many of the cell lines have been characterized as embryonic stem cells by detecting expression of surface antigen markers specific to embryonic stem cells, determining if the cells are pluripotent, and demonstrating that the cells are undifferentiated. A number of scientific publications have described the characterization of human embryonic stem cells. Although the characterization approaches may differ across laboratories, an example of the strategies used can be found in Thomson et al. (1998), Science, 282,1145–1147.
   
   The National Stem Cell Bank and the individual providers of the federally eligible cells are working to make them available to researchers. This includes developing quality control measures to grow and reproduce the cell lines in sufficient numbers, having the administrative structure to receive and process requests, and establishing material transfer agreements with research purchasers. The National Stem Cell Bank and the individual providers of federally eligible cell lines have the most up-to-date information on availability. A list of these sources and contact information is available on the NIH Stem Cell Registry.
   
   
7. What policies govern the use of stem cell lines from WiCell Research Institute?
   
   WiCell has published FAQs About WiCell's Policies on the Use of Its hESC Lines (136k PDF file; get Adobe Reader) to address this question. Click here for more information.
   
   This core also provides hands-on training upon request (see services).
   
   
8. I have experience in mouse ES cell culture? What is the difference between human ES cells and mouse ES cells?
   
   Although human and mouse ES cells share fundamental properties (such as pluripotency) and highly conserved transcriptional networks, they differ in several ways. For example, LIF/STAT3 signaling is critical for mES cell self-renewal, but is dispensable for propagation of undifferentiated hES cells. Together with LIF, BMP4 supports expansion of undifferentiated mES cells, while BMP4 induces trophoblastic differentiation of hES cells. The roles of other signaling pathways in hES cells remain to be fully determined. One thing in common is that primary mouse embryonic fibroblast (pMEFs) are commonly used as feeder cells for maintaining the stocks of both human and mouse embryonic stem cells.
   
   
9. Where can I get feeder cells?
   
   Mouse feeder cells such as pMEFs can be made easily from 13.5-day-old embryos from strains such as CF1 and C57B6 mice. They are also commercially available from several vendors such as GlobalStem and Millipore. Some of them also offer drug resistant (such as hygro). In addition, we also provide immortalized human feeder cells that express 3 drug selection genes (neo, puro and hygro) and a Wnt3a transgene. The human feeders have repeatedly demonstrated the ability to maintain the undifferentiated hES cells.
   
   Ref: Promoting human embryonic stem cell renewal or differentiation by modulating Wnt signal. Cell Research, 17: 62-72.
   
   
10. How do I use feeder-free culture?
    
    It is a fast moving field. We will provide hands-on training or consultation.
    
    
11. Can you help me on directing the differentiation of human ES cells?
    One method is to allow embryoid body formation to initiate differentiation of various lineages. We will provide technical support and consultation for various cell lineages.
    
    
12. I want to express my gene of interest in hES cells stably. I heard it is not easy. Can you help us?
    
    Although plasmid-based stable transfection can be achieved at least in some hES cell lines, it is not efficient (one out of 100,000 colony-forming hES cells). We normally use lentiviral vectors to stably express a transgene (cDNA) or small-hair (sh) RNA in hES cells. We will provide the backbone lentivector and cloning strategy for your project.
    
    With permission of the original hES cell lines provider, we can also provide H1 and H9 hES cell lines that stably express GFP.
    
    Ref. Inducible and reversible transgene expression in human stem cells after efficient and stable gene transfer. Stem Cells, 25: 779-789.
    
    
13. I need some teratoma data for my paper. Can you help?
    
    Yes. We will inject your cells into the immuno-deficient mice as a service. It will take about 10-12 weeks before the teratoma can be palpable. We also provide other sensitive approaches to track the tumor formation if cells express certain reporters (i.e., in vivo imaging).
    
    
14. How do I check for mycoplasma contamination in hES cells?
    
    The core has established the ELISA-based mycoplasma test with high specificity. You can harvest the supernatant of cell culture and send to us. We will report to you within 3 working days.
    
    
15. How do I karyotype my hES cells?
    
    Contact Prenatal Cytogenetics Laboratory, located in Park Building B232.
    Dr. Colyn Cain, Director, 410-955-3476
    
    
16. What other services and costs do you offer?
    
    See the Services section of the website. The cost of karyotyping is set by the Prenatal Cytogenetics Laboratory. When we used last time, the price is about $500 for the whole procedure and reading of up to 20 karyotypes.