新生小鼠小脑颗粒神经元的原代培养与鉴定
Primary culture and identification of cerebellar granule neurons from neonatal mice
方晨昊 1刘辉庆 1夏泽阳 1高俜娉 1鲍南1
作者信息
- 1. 上海交通大学医学院附属上海儿童医学中心神经外科,上海,200127
- 折叠
摘要
目的 通过改进已报道的新生小鼠小脑颗粒神经元培养方法,以获得纯度更高,杂质更少,状态更佳的颗粒神经元.方法 将小鼠小脑组织剪碎,经木瓜蛋白酶消化后得到单细胞悬液,采用Percoll非连续性密度梯度离心法分离出颗粒神经元前体细胞,种植在基质胶包被过的培养皿中并在培养20 h后给予阿糖胞苷(5 μmol/L)处理8h.使用激光共聚焦显微镜及神经元特异性标志物微管相关蛋白2(MAP2)对小脑颗粒神经元细胞进行纯度鉴定.结果 细胞存活率达98.03%±1.048%;神经元纯度为99.71%±0.2%.结论 该方法获取的颗粒神经元存活率和纯度较高,是一种小鼠小脑颗粒神经元体外培养的可靠方法.
Abstract
Objective To enhance the purity and viability of cerebellar granule neurons in cultured newborn mice by refining the previously reported methods.Methods Mouse cerebellar tissue was minced and digested with papain to obtain single-cell suspension.Cerebellar granule neuron precursor cells were separated using the Percoll non-continuous density gradient centrifugation method.These cells were then plated on matrix-coated culture dishes and treated with cytosine arabinoside(5 μmol/L)for 8 hours after 20 hours of culture.Purification of cerebellar granule neurons was assessed using laser confocal microscopy and the neuronal-specific marker,microtubule-associated protein 2(MAP2).Results Cell viability reached 98.03%±1.048%,and neuronal purity was 99.71%±0.2%.Conclusion This method yields high viability and purity of cerebellar granule neurons,providing a reliable approach for in vitro culti-vation of mouse cerebellar granule neurons.
关键词
小鼠/小脑颗粒神经元/原代培养/纯度Key words
Mouse/cerebellar granule neurons/primary culture/purity引用本文复制引用
基金项目
国家自然科学基金(32200781)
国家自然科学基金(81974231)
出版年
2024
NETL
NSTL
万方数据