INTRODUCTIONFast and efficient production of recombinant proteins (r-proteins) remains a major challenge for the academic and biopharmaceutical communities. Such proteins often need to be as pure as possible before any characterization study can begin. Although many types of protein tag are available, histidine is the most popular. Although small-scale immobilized metal-affinity column (IMAC) purification of such proteins (e.g., <500 mL of culture medium) can easily be achieved using gravity chromatography columns, larger volumes can be processed with the aid of automated chromatography systems. This protocol describes an IMAC purification technique for secreted proteins using a cobalt-loaded resin. Preliminary small-scale trials using this technique can be used to determine the production scale that will be needed to provide enough pure material for a given study.