Tube formation assay with endothelial cells on Matrigel

Matrigel (70 μL/well) was added to a 96-well plate and polymerized for 30 min at 37°C. The endothelial cells (3×10⁴ cells) were seeded onto each well of the Matrigel-coated 96-well plate and then incubated in 2% FBSEBM-2 with various concentrations of kaempferol. After incubation for 8 h, the formation of the endothelial cell tubular structures was visualized under an inverted microscope and photographed at 40× magnification. Furthermore, the tube formation was quantified from the calculation of the tube length, normalized, and expressed as a percentage relative to untreated control cells.