诱导多能干细胞衍生的诱导神经元轴突运输活体成像方案。
Protocol for live imaging of axonal transport in iPSC-derived iNeurons.
作者信息
Dou Dan, Holzbaur Erika L F, Boecker C Alexander
机构信息
Department of Physiology, Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA 19104, USA; Neuroscience Graduate Group, University of Pennsylvania Perelman School of Medicine, Philadelphia, PA 19104, USA.
Department of Neurology, University Medical Center Goettingen, 37077 Goettingen, Germany.
出版信息
STAR Protoc. 2025 Mar 21;6(1):103556. doi: 10.1016/j.xpro.2024.103556. Epub 2025 Jan 12.
Studies of human induced pluripotent stem cell (iPSC)-derived neurons promise important insights into neurodegenerative diseases. Here, we present a protocol for live imaging of axonal transport in glutamatergic iPSC-derived neurons (iNeurons). We describe steps for the differentiation of iPSCs into iNeurons via PiggyBac-mediated neurogenin 2 (NGN2) delivery, iNeuron culture and transfection, and the acquisition and analysis of time-lapse images. Our protocol is optimized for the widely available catalog of KOLF2.1J iPSCs with mutations relevant to neurodegenerative diseases but is also applicable to other iPSC lines. For complete details on the use and execution of this protocol, please refer to Dou et al..
对人类诱导多能干细胞（iPSC）衍生神经元的研究有望为神经退行性疾病提供重要见解。在此，我们展示了一种用于对谷氨酸能iPSC衍生神经元（iNeurons）轴突运输进行实时成像的方案。我们描述了通过PiggyBac介导的神经生成素2（NGN2）传递将iPSC分化为iNeurons的步骤、iNeuron培养和转染，以及延时图像的采集和分析。我们的方案针对广泛可用的具有与神经退行性疾病相关突变的KOLF2.1J iPSC目录进行了优化，但也适用于其他iPSC系。有关该方案使用和执行的完整详细信息，请参考窦等人的研究。
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