The elucidation of the biological function of protein complexes is an area of intense research efforts but isolation and analysis of the complexes is challenging as they can be very large and unstable. Superose™ 6 Increase is a new agarose‐based SEC (size exclusion chromatography) medium with a wide separation range, Mr 5000 to 5 000 000 for globular proteins. The wide separation range allows separation of very large proteins, complexes and membrane proteins both for preparative and analytical purposes under mild, non‐dissociative conditions.Separation of a protein involved in structural coloration on Superdex 200 10/300 resulted in co‐elution of two differently sized protein populations. The wider separation range of Superose 6 Increase 10/300 enabled separation of the two populations. Based on elution position, the size of the separated material was estimated to Mr > 700 000Purification of membrane proteins often require addition of expensive detergents in the buffer to keep the proteins stable and thus low consumption of buffer is important. Purification of a membrane protein complex on Superose 6 Increase 5/150 (3 ml bed volume) resulted in good separation from aggregated material and degradation products.

Immobilized metal affinity chromatography (IMAC) purification of secreted histidine‐tagged proteins in eukaryotic cell culture supernatants is often problematic. Incompatibility with the cell culture media appears as stripping of the immobilized metal ions required for binding of target proteins. Due to low target protein concentration in cell culture supernatants, large sample volumes are often needed, aggravating the stripping effect. In this study, purifications of histidine‐tagged proteins were performed using two novel Sepharose™ based IMAC media; Ni Sepharose excel and magnetic His Mag Sepharose excel. Both media have a new type of chelating ligand with exceptionally strong binding of nickel ions. Data showing successful purification of histidine‐tagged proteins from CHO cell culture supernatants will be presented. Furthermore, the characteristics of new media enabled purification of target protein from insect cell culture supernatants. The purification was easily scaled up from 20 μl His Mag Sepharose excel beads to 1 mL pre‐packed columns.

The histidine‐tag is widely used to facilitate purification of recombinant expressed proteins. For immobilized metal‐ion affinity chromatography (IMAC) the most used metal ion is Ni2+. Also Co2+ has been shown useful in obtaining high purity with good yields of histidine‐tagged proteins. Prepacked HiTrap™ TALON® crude column facilitates scalable, fast and easy protein purification of histidine‐tagged proteins. Unclarified samples can be loaded directly to the column reducing the total purification time which helps to ensure high concentrated pure and intact target proteins with minimal degradation. Two proteins (produced in E. coli and P. pastoris) were purified ending up in >90% purity and yields of 90%. An automated purification of a kinase was done by IMAC and gel filtration. SDS‐PAGE analysis showed that the obtained purity of the kinase was above 90% and the amount purified protein was 3 mg. Further, four purification cycles, without cleaning and Co2+‐recharging between the runs, show that the HiTrap TALON crude columns can be used several times without loosing performance. Purity and yield was unchanged