AAT Bioquest

What is the principle of ATP bioluminescence assay?

Posted May 30, 2023


Bioluminescent ATP assays utilize the firefly luciferase enzyme reaction in its mechanism. This reaction uses ATP from viable cells to create photons of light. Living cells are lysed to release ATP for detection, and the reagents with luciferase and the substrate are added to catalyze a two-step reaction. In the first step, luciferin is activated with ATP to produce luciferyl-adenylate and pyrophosphate. In the second step, the luciferyl-adenylate reacts with oxygen to yield oxyluciferin and carbon dioxide. The excited state of oxyluciferin then returns to its ground state with the release of green to yellow bioluminesce light (550-570 nm). The amount of bioluminescence light is measured by a luminometer and expressed in relative light units (RLU). The RLU values are directly proportional to the quantity of ATP. 

Additional resources

Effectiveness of ATP bioluminescence assay for presumptive identification of microorganisms in hospital water sources


Amplite® Renilla Luciferase Reporter Gene Assay Kit *Bright Glow*

PhosphoWorks™ Luminometric ATP Assay Kit *Bright Glow*