诱导多能干细胞
多巴胺能
胚胎干细胞
中脑
神经科学
生物
细胞分化
定向微分
干细胞
细胞生物学
人诱导多能干细胞
移植
医学
多巴胺
中枢神经系统
内科学
遗传学
基因
作者
Fabiano A. Tofoli,Ana Semeano,Ágatha Oliveira-Giacomelli,Maria Carolina Bittencourt Gonçalves,Merari F. R. Ferrari,Lygia V. Pereira,Henning Ulrich
出处
期刊:Methods in molecular biology
日期:2019-01-01
卷期号:: 97-118
被引量:18
标识
DOI:10.1007/978-1-4939-9007-8_8
摘要
The work with midbrain dopaminergic neurons (mDAN) differentiation might seem to be hard. There are about 40 different published protocols for mDAN differentiation, which are eventually modified according to the respective laboratory. In many cases, protocols are not fully described, failing to provide essential tips for researchers starting in the field. Considering that commercial kits produce low mDAN percentages (20-50%), we chose to follow a mix of four main protocols based on Kriks and colleagues' protocol, from which the resulting mDAN were engrafted with success in three different animal models of Parkinson's disease. We present a differential step-by-step methodology for generating mDAN directly from human-induced pluripotent stem cells cultured with E8 medium on Geltrex, without culture on primary mouse embryonic fibroblasts prior to mDAN differentiation, and subsequent exposure of neurons to rock inhibitor during passages for improving cell viability. The protocol described here allows obtaining mDAN with phenotypical and functional characteristics suitable for in vitro modeling, cell transplantation, and drug screening.
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