Strategy for the creation of clinical grade hESC line banks that HLA-match a target population.
Bottom Line: Here, we describe a pre-derivation embryo haplotyping strategy that we developed in order to maximize the efficiency and minimize the costs of establishing banks of clinical grade hESC lines in which human leukocyte antigen (HLA) haplotypes match a significant proportion of the population.Using whole genome amplification followed by medium resolution HLA typing using PCR amplification with sequence-specific primers (PCR-SSP), we have typed the parents, embryos and hESC lines from three families as well as our eight clinical grade hESC lines and shown that this technical approach is rapid, reliable and accurate.By employing this pre-derivation strategy where, based on HLA match, embryos are selected for a GMP route on day 3-4 of development, we would have drastically reduced our cGMP laboratory running costs.
Affiliation: Embryonic Stem Cell Laboratories, Guy's Assisted Conception Unit, Division of Women's Health, King's College School of Medicine, London, UK.Show MeSH
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Mentions: We validated this approach with three couples that underwent PGD for Huntington's disease (HD) in our facility and who had hESC lines derived from affected embryos that were donated for stem cell research (Fig 2; Supporting Information Fig 2). Donor couple #1 had seven affected embryos from which two hESC lines, KCL012 and KCL013, were derived (Ilic et al, 2012). Donor couple #2 had eight affected embryos from which two lines, KCL027 and KCL028, were derived. Donor couple #3 had one affected embryo, from which KCL036 was derived. HLA typing was performed on blood samples from the couples, on the hESC lines and on DNA from the biopsied cells of the embryos. Identical HLA haplotypes were obtained for each embryo-of-origin and hESC line pair, showing that the technical approach to pre-derivation haplotyping is robust and accurate (Fig 2).
Affiliation: Embryonic Stem Cell Laboratories, Guy's Assisted Conception Unit, Division of Women's Health, King's College School of Medicine, London, UK.