Quant it picogreen dsdna quantification assay

HUVECs were suspended in control media at 2.0×10⁴ cells/mL. 0.5 mL of the cell solution was seeded in each well of a 24-well plate. To achieve uniform cell adhesion, cells were allowed to adhere at room temperature for 1 hour before being transferred to the incubator [41 (link)]. 16 hours later, cells were washed twice with PBS and treated with 0.5 mL of either control media with or without peptide drugs or controls. A preliminary dose screening study (data not shown) was conducted to identify concentration at which each “N” peptide induced proliferation, and that concentration was used for both the “N” and “2T” treatments. Media was changed daily, and after 72 hours cells were washed twice with PBS, and lysed via sonication in 1x TE buffer (10 mM Tris, 1 mM EDTA in ddH₂O, pH 7.5). DNA content was quantified using the Quant-iT picoGreen dsDNA quantification assay (Invitrogen). Each plate contained a control media group to account for plate-to-plate variability.