Fig. 1. .

Principle of bioluminescent substrate depletion and product formation assays. Illustrated are the assay principles for protein kinases (Clk4 was used here). (A) ATP-depletion format where a detection reagent containing firefly luciferase is added after the kinase reaction to detect the remaining ATP. (B) ADP-formation assay where following the kinase reaction a reagent that depletes the remaining ATP is added (Reagent I) followed by a reagent that converts the ADP formed in the protein kinase reaction to ATP as well as firefly luciferase that detects the ATP levels (Reagent II). (C) The luminescent response (presented as RLUs, or relative light units) is inversely proportional to the kinase activity in the ATP-depletion format (Kinase-Glo assay) while the luminescent response is directly proportional to kinase activity in the ADP-formation format (ADP-Glo assay).