We describe the steps in detail to expand human neural stem cells, to bank and cryopreserve the stem cells. The methods described in this protocol represent the latest improvement in tissue culture and molecular biology reagents to help identify specific cell lineages. Specific tissue culture media and methods are outlined to characterize neural stem cells and to differentiate the cells to neurons/oligodendrocytes/ astrocytes. The protocols could be modified to obtain specific types of neurons such as spinal cord motoneurons and dopaminergic neurons by incorporating key morphogens during lineage specification.