In the field of proteomics, numerous advanced technologies have evolved that aim to provide the molecular data necessary for an in-depth understanding of biological processes. Protein–protein interactions (PPI) are at the heart of cellular function and a milestone yet to be achieved is the mapping of a complete human interactome. Bioluminescence resonance energy transfer (BRET) has become a popular technique to investigate PPI. As BRET enables the detection of PPI in living cells, problems associated with in vitro biochemical assays can be circumvented, thus making BRET a powerful tool for monitoring interactions of virtually all kinds of protein species.