Derivation of human embryonic stem cells from day-8 blastocysts recovered after three-step in vitro culture

Miodrag Stojkovic, Majlinda Lako, Petra Stojkovic, Rebecca Stewart, Stefan Przyborski, Lyle Armstrong, Jerry Evans, Mary Herbert, Louise Hyslop, Sajjad Ahmad, Alison Murdoch, Tom Strachan

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57 Citations (Scopus)


Human embryonic stem cells (hESCs) have been derived from the inner cell mass (ICM) of day 5-7 blastocysts and hold great promise for research into human developmental biology and the development of cell therapies for the treatment of human diseases. We report here that our novel three-step culture conditions successfully support the development of day-8 human blastocysts, which possess significantly (p <.01) more ICM cells than day-6 blastocysts. Plating of ICMs isolated from day-8 blastocysts resulted in the formation of a colony with hESC morphology from which a new hESC line (hES-NCL1) was derived. Our stem cell line is characterized by the expression of specific cell surface and gene markers: GTCM-2, TG343, TRA1-60, SSEA-4, alkaline phosphatase, OCT-4, NANOG, and REX-1. Cytogenetic analysis of the hESCs revealed that hES-NCL1 line has a normal female (46, XX) karyotype. The pluripotency of the cell line was confirmed by the formation of teratomas after injection into severely combined immunodeficient mice and spontaneous differentiation under in vitro conditions.

Original languageEnglish
Pages (from-to)790-797
Number of pages8
JournalStem Cells
Issue number5
Publication statusPublished - 2004
Externally publishedYes


  • Differentiation
  • Human embryonic stem cells
  • In vitro culture
  • Pluripotency

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