Effect of Y477F ezrin on morphology of AC2M2 cells in 3D Matrigel cultures: Panel A: AC2M2 cell clones expressing pCB6 empty vector or Y477F ezrin (clones A43 and C13) were cultured in 3D Matrigel. Colony morphology was assessed at the times indicated, using phase contrast microscopy. Objective magnification is shown in each panel. Panel B: Table indicates percentages of invasive colonies from each cell clone with branching cellular extensions and protrusions, and of cohesive round colonies with no extensions, respectively. The presence of 5 or more cellular extensions or protrusions per colony was considered positive. Mean percentage of colonies +/- SD (3 wells per group) in each category is shown. N = total number of colonies counted. pCB6 cells formed predominantly invasive colonies with cellular extensions, whereas the majority of Y477F ezrin cells formed round colonies with no extensions. The above differences in colony morphology were significant as determined by one way ANOVA (p< 0.001).