求助：IPS cell 培养以及鉴别诊断的Protocol

stonelee2050

从别处购买了IPS ，怎么样培养，怎么样诱导分化为心肌细胞，怎么样鉴别诊断呢？

xjya

首先你可以看看相关的文献,如果没有可以下载iPS专区的iPS系列经典文献看看.
$ E2 c# R! T+ D* @以下是我们实验室的protocol:(这个对小鼠的ES细胞及其分化同样适用）希望对你有帮助。
2 c0 {6 ~  S$ [. L# z# e9 {! T$ _Cell culture of miPS-CMs
, J. }/ {7 O/ `* }2 y" wThe undifferentiated Nanog-selected iPS cell line N10 was generated from murine embryonic fibroblasts (MEFs) by M.W. and co-workers. Briefly, undifferentiated miPS cells were grown on inactivated MEFs in propagation medium composed of Iscove’s modified Dulbecco’s medium (IMDM) supplemented with 17% fetal calf serum (FCS), 100 U/ml penicillin, 100 _g/ml streptomycin, 1% nonessential amino acids, 0.1 mM _-mercaptoethanol, and 1000 IU/ml LIF. MEFs for propagation of undifferentiated iPS cells were prepared from Him:OF1 outbred mice at embryonic day 14.5 and inactivated by mitomycin C treatment. Undifferentiated miPS cells were passaged every second or third day. Confluent cells were trypsinized into a single cell suspension, centrifuged, and suspended again in IMDM with 20% FCS (without LIF) and maintained on a shaker. After 4 days they were diluted to _1000 embryonic bodies/10-cm dish, and maintained on the shaker until d 7. Then they were plated on 0.1% gelatin-coated culture dishes with 20% IMDM. On day9, beating areas can be detacted.

xjya

不太明白你说的鉴别诊断是什么意思？是如何检测心肌细胞的存在吗？如果是，可以做流式，或是免疫组化检测心肌细胞的特异性标志物比如，a-actinin, tropinin等。

stonelee2050

回复 2# xjya
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' h3 X4 G& n" J5 ~- t9 E    thank you very much!

daviehe

给你推荐一个文献 关于人ips分化成心肌的0 N4 \' Z  Y, A& G; R9 F2 z1 t
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Mouse and human induced pluripotent stem cells as a
. A: `6 [7 q& Z2 ], ]source for multipotent Isl1
cardiovascular
: R% N5 r$ m4 a3 U4 Uprogenitors。作者 Alessandra  Moretti 2 }  \: t9 k  G: D1 w4 S) k6 X. T3 }
期刊 The FASEB Journal