iPS facility

LUMC iPSC core facility

The Board of the LUMC has provided annual support to establish a core facility for the generation and differentiation of human induced pluripotent stem (iPS) cells. The facility is set up and operated jointly by the departments of Anatomy & Embryology and Molecular Cell Biology. Prof. Dr. F. Breedveld, director of the LUMC officially opened the facility on 14 March 2011.

The motivation underlying the core facility start-up is (i) exploiting the unique possibilties within the LUMC for generating patient specific pluripotent stem cell lines for preclinical research; these can give new insights into disease processes and help identify molecular targets through which intervention may help slow down or reverse development of the disease; (ii) providing LUMC research groups access to this technique. (iii) the need to broaden and strengthen further stem cell research within the Regenerative Medicine profile of the LUMC.  

What the facility plans to offer (free of charge for LUMC researchers).

• Isolation and culture of fibroblasts from a skin biopsy or isolation of circulating endothelial cells from peripheral blood (+ freezing stock for banking)
• The generation of iPS cell lines (two patients per disease phenotype) using lentiviral vectors in the absence of GMP
• Expansion of 3-5 patient-specific clones to passage 5 and freezing for banking
• Analysis of the expression of pluripotency markers by immunostaining (OCT4, NANOG, TRA1-81, SSEA4, and SSEA1)
• Evidence of differentiation into three germ layers in vitro (smooth muscle actin-expressing cells (mesoderm), b3 Tubulin expressing cells (ectoderm), and AFP+ endodermal derivatives)
• Training courses for new users at certain times of the year. Researchers accepted for training would be expected to have skills in basic culture/asceptic techniques and preferably experience passage mouse ES cells. In general training will include:
  • Maintanance and passaging of undifferentiated cells in TesR medium without feeders cells (this is far easier than regular culture on feeders although this is in general the best way to maintain long term pluripotency and karyotypic stability)
  • Freeze and thaw of stocks
  • Spontaneous differentiation in embryoid bodies to endodermal, ectodermal and mesodermal derivatives as for the basic characterization.
• A protocol book, with references, relevant papers and current best reagent lists will be provided. Researchers will be able to purchase reagents/culture media used and validated by the facility for use of the generated iPS cell lines in their own labs.
• The facility will make reagents available to labs using the LUMC iPS facility service. Updates of lot numbers of reagents used in the facility will be provided upon request.
• Where possible, assistance will be given to the researchers in searching the literature for robust protocols for their own particular cell of interest.

The facility does not perform karyotyping or teratoma generation in mice (proof of pluripotency in vivo). The researcher can decide to do these later if (one of) the lines received will be in long term use. In addition viral integration sites will not be determined. Removal of floxed transgenes can be requested but will not be included as standard procedure.