The proliferative potential of embryonic stem cells was examined. In contrast to the current concept of t h e finite life-span being the hallmark of normal cells, we have been able to maintain these embryonic stem cells in vitro up to about 250 cumulative doublings with no indication of "crisis" or transformation. These cells could be considered normal on t h e basis of: (1) their apparently normal diploid karyotype, (2) their ability to extensively colonize embryos without causing tumors and developmental anomalies, and (3) their ability to form normal gametes when differentiated into t h e germ-line. These results suggest that embryonic stem cells prior to differentiation into germ and somatic cells are indeed immortal.
Human diploid fibroblasts in culture are endowed with cells were cultured in DMEM with 10% serum, and the finite life-span as first demonstrated by Hayflick feeder cells were prepared by y-irradiation on gelatinand Moorhead (1961). In as much as similar observa-coated dishes. Of various mouse early embryonic stem tions have been made on other types of cells, including cells tested, EK.CP1 was kindly provided by Dr. Evans various stemlike cells, from a variety of species, the at passage (P) 10. The ESC.CD1 series of lines were concept of finite life-span appears applicable to all ver-established as described (