The Raf kinase inhibitor protein (RKIP) belongs to an evolutionarily conserved family of phosphatidylethanolamine-binding proteins (PEBPs), which have important functions as inhibitors of kinase signaling pathways and metastasis. Most notably, RKIP can interrupt signaling through the Ras-Raf-MEK-ERK pathway by dissociating the interaction between Raf-1 and its substrate MEK, highlighting the importance of protein interactions as regulatory interfaces. Furthermore, RKIP was shown to inhibit IkappaB kinases (IKKs) interfering with the activation of nuclear factor kappa B (NFkappaB), and G-protein coupled receptor-kinase 2 (GRK2), impeding receptor downregulation and prolonging signaling. More recently, RKIP has emerged as an important suppressor of metastasis. Here, we review the functions of RKIP and present methods to detect and measure RKIP expression and activity in cells and tissues.