Protocol for quantifying extracellular and intracellular ATP from macrophages upon inflammasome activation using a luciferin-luciferase technique

Extracellular ATP triggers several cellular responses, while intracellular ATP depletion primes cells for death. Here, we present a protocol for the quantification of extracellular and intracellular ATP from bone marrow-derived macrophages upon inflammasome activation using a luciferin-luciferase technique. We describe steps for macrophage isolation, differentiation, and inflammasome activation with real-time ATP measurement. The protocol allows quantitative determination of intracellular ATP and extracellular ATP hydrolysis, with applicability to different cell types. For complete details on the use and execution of this protocol, please refer to Schachter et al.¹.