猪原代神经细胞的体外培养与鉴定

Culture and identification of neural stem cells isolated from pig

为建立猪原代神经细胞体外培养及鉴定方法，本研究选取90日龄潜江黑猪1头，分离猪脑组织，无菌剪碎后采用胰酶消化和机械分离相结合的方法制备神经细胞悬液，进行培养，逐日在倒置相差显微镜下观察，并用间接免疫荧光鉴定。结果显示，培养72h后细胞全部贴壁，并长出突起，随时间增加，突起变长并交错呈网络，培养至15～20d神经细胞状态最佳，随后逐渐老化，神经细胞可生长30d，经鉴定Nestin表达呈阳性。本研究建立的方法适合一般实验室开展猪原代神经细胞的培养与初步鉴定。

To establish a method to isolate culture and identify the neural cells from pig. The neural stem cells were isolated from 90-day pigs. Trypsin digestion and mechanical dissociation were a dopted to conduct monolayer primary culture and were observed in inverted phase microscope dai ly. Identification of neurons was detected with indirect immunofluorescence. The results showed that all cells adhered to the footwall after 72 hours and short nervous processes were found in sub- total neurons,and extended markedly with time and connected into nets. Neurons state were the best when cultured for 15-20 days,then began aging,the grow maintained 30 days. The expression of Nestin was positive identified by immunofluorescence. The method developed in our study was fit for the primary culture and identify of pig neurons in the routine labs.