Abstract
The use of 3D cancer models to study therapy response has a great advantage over conventional 2D cell culture. Here, we present an optimized protocol for breast cancer spheroid and rod-shaped microtissue formation using MicroTissues mold systems. We describe steps for cast formation and treating the 3D models with DNA-damaging agents. We then detail procedures for analyzing the 3D models by whole-mount immunostaining and confocal imaging of fixed samples or with the use of live-cell reporters.