Culture and Isolation of Brain Tumor Initiating Cells

Abstract

This unit describes protocols for the culture and isolation of brain tumor initiating cells (BTIC). The cancer stem cell (CSC) hypothesis suggests that tumors are maintained exclusively by a rare fraction of cells that have stem cell properties. We applied culture conditions and assays originally used for normal neural stem cells (NSCs) in vitro to a variety of brain tumors. The BTIC were isolated by fluorescence activated cell sorting for the neural precursor cell surface marker CD133. Only the CD133⁺ brain tumor fraction contains cells capable of sphere formation and sustained self-renewal in vitro, and tumor initiation in NOD-SCID mouse brains. Therefore, CD133⁺ BTICs satisfy the definition of cancer stem cells in that they are able to generate a replica of the patient's tumor and they exhibit self-renewal ability through serial retransplantation. This established that only a rare subset of brain tumor cells with stem cell properties are tumor-initiating, and, in this unit, we describe their culture and isolation. Curr. Protoc. Stem Cell Biol. 11:3.3.1-3.3.10. © 2009 by John Wiley & Sons, Inc.