人类胚胎干细胞的建系与鉴定
发布时间：2018-11-15 16:28
【摘要】： 本实验室利用临床多余的囊胚经患者知情同意进行人胚胎干细胞(human embryonic stem cells, hES)建系研究,首先用链蛋白酶消化以去除囊胚的透明带,然后直接机械分离出内细胞团,接种到饲养层细胞(feeders)上,在hES细胞培养液中进行培养。一共成功建立了5个hES细胞系,全部来自体外受精(in vitro fertilization,IVF)囊胚,分别命名为J6、J13、J14、J30和J32。 根据hES细胞系公认的鉴定标准,我们对其中新建立的2株细胞系的各项特征进行了检测鉴定。这些hES细胞在体外能长期稳定增殖;表达碱性磷酸酶活性;表达hES细胞特异的标志物Oct-4、SSEA-3、SSEA-4、TRA-1-60、TRA-1-81;在体外能形成包含三个胚层来源分化细胞的类胚体(embryoid body,EB);在免疫缺陷小鼠(SCID mouse)体内能形成包含三胚层来源组织的畸胎瘤;经过细胞的单克隆化,产生两个hES细胞亚系(J6-1和J13-1),并且在体外仍能长期稳定增殖;这两个细胞株的群体倍增时间为31.43小时(J6)和21.38小时(J13)。到目前为止,这5个细胞系在体外已经分别培养485(J6)、478(J13)、259(J14)、120(J30)、121(J32)天,培养代数达46(J6)、55(J13)、19(J14)、25(J30)、24(J32)。
[Abstract]:Human embryonic stem cells (human embryonic stem cells, hES) were established by using clinical superfluous blastocysts with the informed consent of patients. Firstly, the blastocysts were digested with strepsin to remove the pellucida of blastocysts, and then the inner cell masses were mechanically isolated. (feeders) was inoculated into feeder layer cells and cultured in hES cell culture medium. Five hES cell lines, all from (in vitro fertilization,IVF blastocyst, were successfully established and named J6 J13, J14, J30 and J32, respectively. According to the generally accepted criteria for the identification of hES cell lines, we detected and identified the characteristics of two newly established cell lines. These hES cells can proliferate steadily for a long time in vitro, express alkaline phosphatase activity and express Oct-4,SSEA-3,SSEA-4,TRA-1-60,TRA-1-81;, a specific marker of hES cells. In vitro, embryoid bodies (embryoid body,EB) containing three embryo-derived differentiated cells were formed in vitro, teratoma containing three-layer derived tissue was formed in (SCID mouse) of immunodeficient mice. Two hES cell sublines (J6-1 and J13-1) were produced by cell monoclonal transformation, and the population doubling time of these two cell lines was 31.43 hours (J6) and 21.38 hours (J13). So far, 485 (J6), 478 (J13), 259 (J14), 120 (J30), 121 (J32) days have been cultured in vitro, and the cultured algebras are 46 (J6), 55 (J13), 19 (J14), 25 (J30), 24 (J32).
【学位授予单位】：内蒙古农业大学
【学位级别】：硕士
【学位授予年份】：2009
【分类号】：R329
本文编号：2333813
[Abstract]:Human embryonic stem cells (human embryonic stem cells, hES) were established by using clinical superfluous blastocysts with the informed consent of patients. Firstly, the blastocysts were digested with strepsin to remove the pellucida of blastocysts, and then the inner cell masses were mechanically isolated. (feeders) was inoculated into feeder layer cells and cultured in hES cell culture medium. Five hES cell lines, all from (in vitro fertilization,IVF blastocyst, were successfully established and named J6 J13, J14, J30 and J32, respectively. According to the generally accepted criteria for the identification of hES cell lines, we detected and identified the characteristics of two newly established cell lines. These hES cells can proliferate steadily for a long time in vitro, express alkaline phosphatase activity and express Oct-4,SSEA-3,SSEA-4,TRA-1-60,TRA-1-81;, a specific marker of hES cells. In vitro, embryoid bodies (embryoid body,EB) containing three embryo-derived differentiated cells were formed in vitro, teratoma containing three-layer derived tissue was formed in (SCID mouse) of immunodeficient mice. Two hES cell sublines (J6-1 and J13-1) were produced by cell monoclonal transformation, and the population doubling time of these two cell lines was 31.43 hours (J6) and 21.38 hours (J13). So far, 485 (J6), 478 (J13), 259 (J14), 120 (J30), 121 (J32) days have been cultured in vitro, and the cultured algebras are 46 (J6), 55 (J13), 19 (J14), 25 (J30), 24 (J32).
【学位授予单位】：内蒙古农业大学
【学位级别】：硕士
【学位授予年份】：2009
【分类号】：R329
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相关期刊论文 前3条
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