HUVEC cell proliferation assay

HUVEC cells were plated in a 96-well plate at 5,000 cells per well, in triplicate. Once the cells had adhered to the well walls, the medium was replaced with CM from transfected MG63 or U2-OS cells. An MTT assay was performed at 0 and 48 h from the addition of CM. Cells were incubated with MTT for 4 h, then formazan was dissolved with 0.1% dimethyl sulfoxide. The absorbance of the wells was then measured at 490 nm. The proliferation rate was calculated as OD at 48 h/OD at 0 h.