A gene coding for an anti-apoptotic 30K protein originating from the silkworm, Bombyx mori, was cloned into the vector pET 22b(+) to produce this protein in Escherichia coli. The 30K gene was fused to a C-terminal His-tag to facilitate the separation process. The 30K protein was expressed in the form of an inclusion body. This was denatured, purified by single-step immobilized metal ion affinity chromatography (IMAC), and refolded by two different methods, i.e. on-column refolding and refolding by dilution after column chromatography. The latter method resulted in the higher separation yield of the 30K protein. The purified 30K protein effectively inhibited insect cell (Sf9) and mammalian cell (HeLa) apoptosis by addition to culture medium. (C) 2003 Elsevier Inc. All rights reserved.