Preparation of dissociated mouse cortical neuron cultures.
Preparation of dissociated mouse cortical neuron cultures.
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DOI：
10.3791/562
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发表时间：
2007-01-01
期刊：
影响因子：
--
通讯作者：
Smith, Martin A
中科院分区：
文献类型：
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作者：
Hilgenberg, Lutz G W;Smith, Martin A
本视频将指导您完成从胚胎晚期和出生后早期小鼠大脑中产生皮质神经元培养物的过程。这些培养物可用于多种应用，包括免疫细胞化学、生物化学、电生理学、钙和钠成像、蛋白质和/或RNA分离。这些培养物还提供了一个平台，以研究携带晚期胚胎或出生后致命基因突变的转基因动物的神经元发育。这个过程相对简单，需要一些组织培养技术的经验，如果你准备好了，不应该超过两到三个小时。从丘脑-皮质纤维束仔细分离皮质外皮将减少不需要的非神经元细胞的数量。为了增加神经元细胞的产量，在酶孵育步骤后轻轻研磨皮质组织的碎片。这是必要的，因为它可以防止对细胞的不必要的损伤和神经元细胞的过早死亡。由于这些培养物是在没有神经胶质饲养细胞的情况下维持的，它们还提供了一个额外的优势，即培养富含神经元的培养物。
This video will guide you through the process for generating cortical neuronal cultures from late embryo and early postnatal mouse brain. These cultures can be used for a variety of applications including immunocytochemistry, biochemistry, electrophysiology, calcium and sodium imaging, protein and/or RNA isolation. These cultures also provide a platform to study the neuronal development of transgenic animals that carry a late embryonic or postnatal lethal gene mutation. The procedure is relatively straight forward, requires some experience in tissue culture technique and should not take longer than two to three hours if you are properly prepared. Careful separation of the cortical rind from the thalamo-cortical fiber tract will reduce the number of unwanted non-neuronal cells. To increase yields of neuronal cells triturate the pieces of the cortical tissue gently after the enzyme incubation step. This is imperative as it prevents unnecessary injury to cells and premature neuronal cell death. Since these cultures are maintained in the absence of glia feeder cells, they also offer an added advantage of growing cultures enriched in neurons.