The utility and limitations of neurosphere assay, CD133 immunophenotyping and side population assay in glioma stem cell research

Abstract

The newly proposed glioma stem cell (GSC) hypothesis may re-model the way we diagnose and treat the tumor, which highlights the need for a complete knowledge on the genetic and epigenetic "blueprints" of GSCs. To identify the true "stemness" signatures, pure GSC populations are primarily needed. Reliable in vitro methods enriching for GSCs and thereby identifying the key stem-like characteristics constitute the preliminary step forward. We discuss in this review the current widely used methods for enriching and isolating GSCs, namely neurosphere assay, CD133 Immunophenotyping and side population assay, and detail their limitations and potential pitfalls that could complicate interpretation of corresponding results.

Figure 1

Schematic illustration of the comparative constitutions of the representative cell populations. Tumor spheres, CD133+ cells and side population cells are represented as green, blue and yellow ellipses, respectively, and constitute only small proportions of the total tumor cells, represented by the gray area. None of the three populations corresponds to the entire glioma stem cell (GSC) pool, represented by the red ellipse in the center of the bulk tumor cells. Each population of cells comprises a heterogeneous collection of cells moderately enriched for GSCs and marginally overlapping with each other. A combination of methods might produce purer cell fractions than any one individual method, as indicated by the dashed core. Additional assays or markers need to be developed to define the GSCs beyond the reach of each of the three ellipses.