The gaseous hormone ethylene regulates multiple aspects of plant growth and development, as well as responses to biotic and abiotic factors. The copper transporter RAN1 mediates the delivery of copper ions to ETR1 and is crucial for proper ethylene receptor function and thus proper ethylene signaling. Moreover, the central regulator of copper homeostasis, SPL7 interacts with RAN1 at the endoplasmic reticulum to modulate ethylene signaling. Here, we present a detailed protocol of luciferase complementation imaging (LCI), a bioluminescence technique, to examine the interaction between membrane-bound proteins, such as SPL7 and RAN1, in ethylene signaling.