Unlike most cell-based BrdU assay kits that can only measure qualitative differences in BrdU incorporation, the CytoSelect™ BrdU Competitive ELISA Kit quantifies BrdU incorporated into cellular DNA. The unknown DNA samples or standards are first added to a microplate preadsorbed with a BrdU conjugate. After a brief incubation an anti-BrdU monoclonal antibody is added, followed by an HRP-conjugated secondary antibody. The BrdU in solution and the BrdU coated on the plate compete for binding of the antibody. If no BrdU is in solution, all of the antibody will bind to the plate resulting in a high signal. High levels of BrdU in solution will bind the antibody and be washed away, resulting in a low signal.
Assay Principle for the CytoSelect™ BrdU Cell Proliferation ELISA Kit.
Serum Stimulation of Proliferation in HEK 293 Cells. Cells were plated overnight at 37ºC. Cells were then incubated in the presence or absence of 10% FBS for 24 hours, followed by treatment with 10 µM BrdU for 1 or 3 hours. Cells were tested for BrdU incorporation according to the assay protocol.