Measuring Odorant Receptor Activation using a Real-Time Cyclic Adenosine Monophosphate Assay

Take a multiwell plate containing transfected mammalian cells.

These cells express the olfactory neuron's odorant receptor (OR), a G-protein-coupled receptor, along with a cAMP biosensor containing a cAMP binding domain fused to a mutant luciferase.

Remove the medium and wash with buffer to remove the residual medium.

Add a buffer containing a cAMP biosensor substrate. Incubate to facilitate substrate uptake into the cells.

Using a chemiluminescence plate reader, measure the basal luminescence of the cells.

Add increasing concentrations of the test odorant to the wells, leaving one well untreated as a control.

The odorant binds to the OR and triggers a signaling cascade that produces cAMP. 

This cAMP binds to the biosensor's binding domain, activating luciferase, which interacts with the substrate to emit light.

Measure the luminescence in real time. An increase in luminescence with higher odorant concentrations indicates OR activation by the odorant.