- Human neural stem cells (hNSCs) are a valuable tool in brain cancer research since they are used as a normal control for multiple assays, mainly pertaining to toxicity. Here, we present a protocol to safely and successfully derive and culture hNSCs in vitro from human embryonic brain tissue. We describe the steps to dissociate embryonic brain tissue and culture hNSCs, followed by the procedure to expand hNSCs. These cells can be used for downstream applications including RNA-seq and omics studies. For complete details on the use and execution of this protocol, please refer to Venugopal et al. (2012b), Bakhshinyan et al. (2019), and Venugopal et al. (2012a).