Human neural stem cells (hNSCs) and human induced pluripotent stem cells (hiPSCs) have been the primary focuses in basic science and translational research as well as in investigative clinical applications. Therefore, the capability to perform reliable derivation, effective expansion, and long-term maintenance of uncommitted hNSCs and hiPSCs and their targeted phenotypic differentiations through applying chemically and biologically defined medium in vitro is essential for expanding and enriching the fundamental and technological capacities of stem cell biology and regenerative medicine. In this chapter, we systematically summarized a set of protocols and unique procedures that have been developed in the laboratories of Prof. Teng and his collaborators. These regimens have been, over the years, reproducibly and productively used to derive, propagate, maintain, and differentiate hNSCs, including those derived from hiPSCs. We emphasize the multimodal methodologies that were pioneered and established in our laboratories for characterizing functional multipotency of stem cells and its value in basic science as well as translational biomedical studies.