A general Bioluminescence Resonance Energy Transfer (BRET) protocol to measure and analyze protein interactions in mammalian cells

Bioluminescence resonance energy transfer (BRET) allows to quantitate protein interactions in intact cells. Here we provide a step-by-step protocol for measuring BRET due to transient interactions of oncogenic K-RasG12V in plasma membrane nanoclusters of HEK293-EBNA cells. We describe how to seed, transfect and replate cells, followed by their preparation for BRET-measurements on a microplate reader and detailed data analysis steps. For details on how to apply this protocol, please refer to Steffen et al., 2024 1. Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=200 SRC="FIGDIR/small/602189v1_ufig1.gif" ALT="Figure 1"> View larger version (36K): org.highwire.dtl.DTLVardef@152e8c2org.highwire.dtl.DTLVardef@2f2aacorg.highwire.dtl.DTLVardef@9a8e9borg.highwire.dtl.DTLVardef@108833f_HPS_FORMAT_FIGEXP M_FIG C_FIG