Experiments for the determination of luciferase

Summary

Detection of Firefly Luciferase Activity by Using Luciferin as a Substrate

Principle

Luciferin 4-monooxygenase (ATP-hydrolyzed) extracted from firefly, Photinus Pyralis.

Fluorescein + ATP + O2 → oxygen and worm fluorescein + PPi + H2O + light

The optical density I is related to the Michaelis-Menten equation

where H is the Planck constant and v is the frequency of emitted light. When the ATP concentration is very low ([ATP] < < Km), the Michaelis-Menten equation simplifies to:

I = Constant × [ATP]

Constant = V/Km. optical density is proportional to [ATP].

Appliance

cell-based assay

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