sDll4 induces sprouting in endothelial cell spheroids in vitro. (A) HUVEC spheroids were cultured on Matrigel in growth factor–deficient conditions in triplicates with either sDll4 or VEGF for 24 to 72 hours. Shown are representative pictures using sDll4-His from triplicate wells repeated twice. (B) Quantitative analysis for vascular area is shown (Bioquant Image Analysis; mean ± SEM from triplicate wells in 2 repetition experiments). Similar results were seen with sDll4-Fc (data not shown). Photomicrographs were taken using a Nikon Coolpix 5000 camera and a Carl Zeiss Invertoskop microscope with a Nikon Plan Fluor ∞, 0.17, 4×/0.12 NA objective and 10× eyepiece and processed with Image-Pro Plus 6.0. Experiments were repeated with similar results. (B) *P < .05 compared to no growth factor.