Journal
BIOTECHNIQUES
Volume 38, Issue 6, Pages 879-+Publisher
EATON PUBLISHING CO
DOI: 10.2144/05386ST01
Keywords
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Funding
- NIDDK NIH HHS [DK061844] Funding Source: Medline
- NIGMS NIH HHS [GM069983-01] Funding Source: Medline
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A significant obstacle to using human embryonic stem cells (hESCs) arises from extremely poor survival associated with freezing, typically, in the range of 1%. This report describes a slow controlled-rate freezing technique commonly used for mammalian embryo cryopreservation. Using a combination of surviving colony number and colony diameter survival was determined relative to untreated hESCs. Using a dimethyl sulfoxide (DMSO) cryoprotectant and either a homemade controlled-ratefreezing device or a commercial freezing device, survival rates of 20%-80% were obtained. To achieve the highest levels of survival, the critical factors were an ice crystal seed (at -7 degrees to -10 degrees C), afreeze rate between 0.3 degrees and 1.8 degrees C/min, and a rapid thaw rate using room temperature water. Slow controlled-rate cooling allows a rapid, simple, and reproducible means of cryopreserving hESCs.
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