Subcellular localization of Survivin ΔEx3 in HUVE cells. HUVE cells were seeded on glass coverslips and immunostained with antibodies against Survivin ΔEx3, alpha tubulin, or GM130, or the mitochondrial-specific dye Mitotracker, as described in “Materials and methods.” (A) In dividing cells, Survivin ΔEx3 (red) was localized in the cytoplasm, and did not colocalize with alpha-tubulin (green) at any stage of mitosis (note absence from the midbody/cleavage furrow [arrows]). (B) In resting cells, Survivin ΔEx3 (green) partially colocalized with the Golgi apparatus marker GM130 (red; colocalization resulting in yellow pixels, arrows), but did not colocalize with Mitotracker (red). In a number of cells, Survivin ΔEx3 also localized to the actin cytoskeleton (green, arrow) as well as to the spreading edge of the cell, resembling a membrane ruffle (green, arrow). Nuclear localization was also observed in a minority of cells (arrow; nucleus was pseudocolored red for better contrast, and in all other images the nucleus is depicted in blue from Hoechst staining). Scale bar represents 20 μm.