EphA2 silencing reduces cell invasive growth in 3D collagen. (A) The breast carcinoma cells were embedded in 3D collagen as a single cell suspension followed by a 4-d culture. Confocal micrographs show EphA2 and filamentous actin (phalloidin) in representative cell colonies. Arrowheads indicate round single cells with prominent surface and intracellular EphA2. See Fig. S1 G for details. (B and C) Quantitative assessment of single-cell invasion (B) and cell shape (C; error bars indicate mean ± SEM; three collagen preparations per cell line). (D) Confocal micrographs show representative colonies of the phalloidin-stained (red) invasive cells after MT1-MMP and EphA2 knockdown. Relative colony size is indicated in each micrograph (mean ± SEM; six collagen preparations per cell line). Broken lines define individual colonies. P-values were determined with a Mann–Whitney U test (B–D). Bars, 50 µm.