The brain tumor stem cell (BTSC) hypothesis is based on the premise that there is a subpopulation of cells within tumors with tumorigenic and pluripotent properties. BTSC are believed to be responsible for both the initiation of brain tumors and their resistance to current therapeutic modalities. This new paradigm stresses the need for adequate techniques to culture and characterize this special population of cells. Furthermore, the use of different cell migration assays offers the possibility to evaluate the processes involved in glioma metastasis. In this chapter, we summarize a method to culture, analyze the cellular characteristics, and study the invasion of BTSCs using a neurosphere assay, cryostat sectioning, and human organotypic brain cortex migration assay, respectively.