Effective cryopreservation of human embryonic stem cells by the open pulled straw vitrification method.
BACKGROUND
Human embryonic stem (ES) cells originate from the inner cell mass of the blastocyst, and retain in culture the properties of pluripotent cells of the early embryo. The study aim was to determine whether the open pulled straw (OPS) vitrification method, which is highly effective for the cryopreservation of embryos, might be also efficient for human ES cells.


RESULTS
All human ES cell clumps that were vitrified by the OPS method could be recovered upon thawing, and gave rise to ES cell colonies after plating. Vitrified colonies were significantly smaller and showed an increased level of background differentiation compared with control colonies. However, these unwanted effects could be overcome by additional cultivation of the colonies for 1 and 2 days respectively. The vitrified human ES cells were cultivated for prolonged periods and retained the properties of pluripotent cells, including a normal karyotype, expression of the transcription factor Oct-4 and surface markers that are characteristic to human ES cells. When grafted into SCID mice, the vitrified cells gave rise to teratomas containing derivatives of all three embryonic germ layers.


CONCLUSIONS
Vitrification by the OPS method is reliable and effective for the cryopreservation of human pluripotent embryonic stem cells.
Reubinoff
B E
BE
Department of Obstetrics and Gynecology, Goldyn Savad Institute of Gyne Therapy, Hadassah University Hospital, Ein-Kerem, Jerusalem 91120, Israel. reubinof@md2.huji.ac.il
Pera
M F
MF
Vajta
G
G
Trounson
A O
AO
D000818:Animals; D002454:Cell Differentiation; D002455:Cell Division; D002460:Cell Line; D002478:Cells, Cultured; D015925:Cryopreservation / Q000379:methods*; D004268:DNA-Binding Proteins / Q000378:metabolism; D004622:Embryo, Mammalian / Q000166:cytology*; D006801:Humans; D007621:Karyotyping; D008297:Male; D051379:Mice; D016513:Mice, SCID; D050814:Octamer Transcription Factor-3; D012016:Reference Values; D033581:Stem Cell Transplantation; D013234:Stem Cells / Q000166:cytology / Q000502:physiology; D013724:Teratoma / Q000209:etiology / Q000473:pathology; D013736:Testicular Neoplasms / Q000209:etiology / Q000473:pathology; D013997:Time Factors; D014157:Transcription Factors / Q000378:metabolism; D014183:Transplantation, Heterologous
D004268:DNA-Binding Proteins; D050814:Octamer Transcription Factor-3; C494173:POU5F1 protein, human; C494174:Pou5f1 protein, mouse; D014157:Transcription Factors