期刊
PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA
卷 105, 期 36, 页码 13409-13414出版社
NATL ACAD SCIENCES
DOI: 10.1073/pnas.0806136105
关键词
defined serum-free culture; feeder-free
资金
- Medical Research Council, U.K
- Juvenile Diabetes Research Foundation
- Kanagawa Dental College
- Medical Research Council Stem Cell Career Development Fellowship
- MRC [G0801059, G0500491, G0300273, G0700785] Funding Source: UKRI
- Medical Research Council [G0500491, G0801059, G0300273, G0700785] Funding Source: researchfish
A major limitation in developing applications for the use of human embryonic stem cells (HESCs) is our lack of knowledge of their responses to specific cues that control self-renewal, differentiation, and lineage selection. HESCs are most commonly maintained on inactivated mouse embryonic fibroblast feeders in medium supplemented with FCS, or proprietary replacements such as knockout serum-replacement together with FGF-2. These undefined culture conditions hamper analysis of the mechanisms that control HESC behavior. We have now developed a defined serum-free medium, hESF9, for the culture of HESCs on a type I-collagen substrate without feeders. In contrast to other reported media for the culture of HESCs, this medium has a lower osmolarity (292 mosmol/liter), L-ascorbic acid-2-phosphate (0.1 mu g/ml), and heparin. Insulin, transferrin, albumin conjugated with oleic acid, and FGF-2 (10 ng/ml) were the only protein components. Further, we found that HESCs would proliferate in the absence of exogenous FGF-2 if heparin was also present. However, their growth was enhanced by the addition of FGF-2 up to 10 ng/ml although higher concentrations were deleterious in the presence of heparin.
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