人多巴胺神经元细胞-RFP(RFP-HDaNCs)

人类多巴胺能神经元细胞是从胎儿脑组织中分离出来的，这些组织是从授权采购和分发研究组织的机构获得的。多巴胺能神经元细胞选择在无酪氨酸的培养基中，辅以混合生长因子。用免疫细胞化学方法检测细胞中酪氨酸羟化酶（TH）的表达。将表达RFP的慢病毒感染人多巴胺能神经元细胞（RFP-HDaNCs）后，用嘌呤霉素筛选出表达RFP的人多巴胺能神经元细胞（RFP-HDaNCs）。在P2代细胞中，每小瓶提供5x10[5]个细胞。细胞在常规的组织培养皿中生长，在合流处以1:2的分裂率传代培养。我们建议不要使用超过4代的细胞。4代后，酪氨酸羟化酶阳性细胞的数量可能会显著减少，因为它们没有保持在选择性培养基中，只允许表达酪氨酸羟化酶的细胞生长。

Human dopaminergic neuronal cells are isolated from fetal brain tissue obtained from agencies authorized to procure and distribute tissues for research. Dopaminergic neuronal cells are selected in tyrosine free medium supplemented with a mixture of growth factors. Actively growing population of cells are tested for tyrosine hydroxylase (TH) expression by immunocytochemistry. RFP-Expressing Human dopaminergic neuronal cells (RFP-HDaNCs) are selected by puromycin after HDaNCs are infected with lentiviruses expressing RFP. Cells are provided with more than 5x10[5] cells/ vial at passage 2. The cells are grown in regular tissue culture dishes and subcultured at a split ratio of 1 to 2 at confluence. We recommend not to use the cells beyond 4 passages. There may be a significant decrease in the number of tyrosine hydroxylase positive cells after 4 passages as they are not maintained in selective medium to allow the growth of only cells expressing tyrosine hydroxylase.

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