Development of a new coating substrate for human embryonic stem cell culture
Received:October 08, 2013  
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DOI:10.3969/j.issn.1005-4847.2014.03.018
KeyWord:Human embryonic stem cell;Mouse embryonic fibroblast;Methanol fixation;Mouse embryonic fibroblast, MEF-protein complex
                 
AuthorInstitution
朱海林 四川大学生物治疗国家重点实验室, 成都 ;江阴司特易生物技术有限公司, 江苏 江阴
巩慧 江阴司特易生物技术有限公司, 江苏 江阴
周向雅 江阴司特易生物技术有限公司, 江苏 江阴
杨金亮 四川大学生物治疗国家重点实验室, 成都
魏于全 四川大学生物治疗国家重点实验室, 成都
陈慧敏 四川大学生物治疗国家重点实验室, 成都 ;江阴司特易生物技术有限公司, 江苏 江阴
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Abstract:
      Objective To reduce the animal component contamination for human embryonic stem cells (hESCs) and to simplify hESCs culture process, we develop a new coating substrate which can support the hESCs growth without differentiation, and is easy to store and use. Methods Mouse embryonic fibroblasts(MEF)were fixed on the surface of plate by methanol. hESCs were cultured on this new substrate and were passaged every 5 to 6 days. After 10 passages, we checked the cell morphology, alkaline phosphatase expression, embryonic specific markers and the differentiation ability in vitro. Results After 10 passages, the hESCs grew well on this new substrate and maintained the typical hESCs morphology. Alkaline phosphatase staining was positive. Immunofluorescence staining showed that the expressions of Oct4, SSEA4, Tra-1-60 were positive. The cells formed embryoid body in vitro. Conclusions This methanol-fixed MEF substrate can support the growth of undifferentiated hESCs. The coating material can be produced in large scale and stored for a long time. It provides a new and relatively easy way to amplify hESCs.
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