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EXPERIMENTAL PROTOCOLS FOR EMBRYONIC STEM CELL RESEARCH |
a Department of Obstetrics and Gynecology,
b Institute of Reproductive Medicine and Population, Medical Research Center, College of Medicine, Seoul National University, Korea;
c Central Research Institute, Sam Jin Pharm. Co., Ltd., Hwasung, Korea;
d R&D Center, Jeil Pharm. Co., Ltd., Yongin, Korea;
e Department of Physiology, Yonsei University College of Medicine, Seoul, Korea
Key Words. Human embryonic stem cells • Stem cell expansion • Mechanical transfer • Enzymatic transfer • Cell line maintenance
Correspondence: Shin Yong Moon, M.D., Ph.D., Department of Obstetrics and Gynecology, College of Medicine, Seoul National University, 28 Yongon-dong, Chongno-gu, Seoul 110-744, Korea. Telephone: 82-2-2072-2384; Fax: 82-2-3672-7601; e-mail: shmoon{at}plaza.snu.ac.kr; and Dong-Wook Kim, Ph.D., Department of Physiology, Yonsei University College of Medicine, 134 Shinchon-dong, Seodaemun-gu, Seoul 120-752, Korea. Telephone: 82-2-2228-1703; Fax: 82-2-393-0203; e-mail: dwkim2{at}yumc.yonsei.ac.kr
The manipulation of human embryonic stem cells (hESCs) requires refined skills. Here we introduce both mechanical and enzymatic transfer methods for hESCs depending on experimental purpose. We use the mechanical transfer method for maintenance of hESC lines. Although the method is laborious and time-consuming, the technique permits efficient transfer of undifferentiated hESCs and results in similar clump sizes. We implement the enzymatic transfer method when we need the bulk production of cells for various experiments. The enzyme-treated expansion rapidly produces greater amounts of hESCs within a limited time frame. However, the cell clumps vary in size, and there is a probability that both the differentiated and undifferentiated cells will be transferred. In cases in which there are differentiated colonies, the combination of two methods allows mass production of hESCs by excluding differentiated colonies from passage by manual selection before enzyme treatment.
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