Production and Purification of Recombinant Human CLU

Recombinant forms of human CLU, LCN1 and TIMP1 were purchased from R&D Systems (Minneapolis, MN, USA). Natural CLU was purified by immunoaffinity chromatography of plasma prepared from human blood, as described [52 (link)]. A novel recombinant form of human CLU was developed starting from an expression construct previously described [48 (link)]. The novel expressed protein, rhCLU-αC-H2S, incorporates a hexahistidine and twin-strep tags at the C-terminus of the CLU α-chain, a modification to the original twin-strep tagged construct [48 (link)]. The full-length cDNA, carried in the plasmid pRc CMV (Invitrogen), was transiently expressed in human cells (MEXi-293E, IBI Lifesciences, Gottingen, Germany) and the secreted protein was purified from the cell culture medium according to previously published methods [48 (link)].