Figure 1.

Schematic illustrating strategy of drug treatments in sphere-formation assay. After isolating single cell suspension from PCa tissues or PCa cell lines, drugs can be added to every generation in the sphere-formation assay (A), or drugs can be added to the first generation of spheres only (G1D0) and then spheres can be serially propagated to investigate whether the effect is permanent or reversal (B), or spheres can be formed and serially passaged so that you have extensively grew and enriched for stem cells, and then drugs are added at G5D0 to potentially target those cells (C). The sphere-formation efficiency has to be calculated for each generation to assess self-renewal ability of sphere-forming cells. At each generation, spheres could be processed for immune-histochemical analysis to check for differentiation markers, or proteins and total RNA could also be extracted to assess for differentiation.