DUBs are involved in various pathways and signalling networks by pairing with E3 ubiquitin ligases in protein complexes. In order to test whether OTUD5 associates with an E3 ubiquitin ligase to regulate cell proliferation, the stable Flag-OTUD5/Hep3B cell line was established by transfecting Hep3B cells with plasmids expressing pCIN4-Flag-OTUD5. The cellular factors interacting with OTUD5 were purified by anti-Flag antibody-conjugated M2 beads.