目的 改良并比较机械分离法、胰蛋白酶消化法和木瓜蛋白酶消化法在大鼠皮质神经元培养中的优缺点,为科技工作者根据各自条件选择合适的培养方法提供理论依据。方法 采用胎龄为16~18 d 的SD大鼠胚胎,分别用机械分离、胰酶消化以及木瓜蛋白酶消化三种方法对SD胎鼠皮层神经元进行培养,并对不同培养时间的神经元进行形态学观察和免疫荧光显微镜纯度鉴定。结果 三种方法均能成功培养出杂质少且纯度高的皮层神经元,其在不同生长阶段具有典型的形态学特征。经NSE免疫荧光染色,神经元纯度分别在96.28%,95.63% 及97.34%,三组间无统计学差异(P>0.05)。结论 三种方法均成功培养出杂质量较少且纯度高的皮层神经元,均可作为神经元体外培养的良好实验模型,且三种培养方法都有其不同的特点。
Objective To compare three methods for culture of fetal cortical neurons of SD rats and find out the suitable culture conditions of fetal cortical neurons in vitro. Methods The cortex of 16-18-day embryonic rat was used for culture in this study. Mechanical dissociation, trypsin digestion and papain digestion were applied respectively to the neuron culture. The morphological characteristics of neuronal cells at different time points were observed and neuron purity was identified by immunofluorescence staining assay. Results High purity of the fetal rat cortical neurons was successfully achieved by all the three culture methods, and each had distinct morphological characteristics at different time points. The purity of neurons was 96.28%, 95.63% and 97.34%, respectively, with no significant differences among the three groups (P>0.05).Conclusions The three culture methods are improved in our study. Stable neurons with high purity can be obtained by all the three methods respectively, and each of these methods has distinct characteristics.