The histidine‐tag is commonly used to facilitate purification of recombinant proteins. For immobilized metal‐ion affinity chromatography (IMAC) the most used metal ion is nickel. However, also cobalt has been shown useful obtaining high purity with good yield of his‐tagged proteins in IMAC applications.We have designed three formats containing cobalt IMAC media, convenient for rapid and simple small‐scale purification and screening of his‐tagged proteins. The formats, a gravity flow column, a spin column and a 96‐well plate, all allow efficient parallel purification of proteins in μg to mg scale within 40 minutes using fast and easy protocols to help ensure pure target proteins. His‐tagged green fluorescent protein expressed in E. coli was used as model protein to demonstrate the functionality of the formats. The results show that recovery and purity is > 80 % using all formats. The repeatability has proven to be high for all formats with < 10 % difference from average for each run. A comparison between cobalt and nickel IMAC media was performed and our products containing cobalt IMAC media show higher purity compared to products containing nickel IMAC media while still maintaining a high yield.

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

The success of protein production and subsequent analysis are dependent on the expression levels, solubility and purification of the protein. Maltose Binding Protein (MBP) is often used as an affinity fusion tag. Several studies have shown that MBP‐tagged proteins have high solubility compared to other tagged proteins.The new MBPTrap™ HP consists of an affinity medium, Dextrin Sepharose™ High Performance, packed in 1‐ml and 5‐ml HiTrap™ columns. The medium has high specificity and capacity. Additionally, it can withstand harsh Cleaning In Place (CIP) procedures, like sodium hydroxide, without losing binding capacity. The HiTrap format enables automated protocols to be used on ÄKTAdesign™ systems as well as with syringe or laboratory pump.In this work, MBP‐tagged proteins from E. coli were purified on prepacked 1‐ml or 5‐ml MBPTrap HP columns. The purity of the eluted target proteins was greater than 90 %. A stability study was performed, six purification runs each followed by a CIP run (0.5 M NaOH) were made on the same column. The results show no significant change in yield or purity. The results of an automated two step purification on ÄKTAxpress™ is shown. The affinity chromatography step on MBPTrap HP was followed by a gel filtration step.In summary, the results show that the prepacked MBPTrap HP column has high binding capacity and the eluted target protein is very pure. Furthermore, the medium is stable in 0.5 M NaOH.

Water-soluble hemicelluloses were extracted from spruce chips by heat-fractionation using microwave treatment. A screening of conditions (pH, temperature and residence time) was performed for the extraction of O-acetyl-galactoglucomannan (AcGGM). The yield and the average molecular weight of the extracted mannan were analysed using HPLC, size-exclusion chromatography (SEC) and mass-spectrometry. The pH during heat fractionation influenced the yield and the structure of AcGGM. The highest yield (78%) of AcGGM (average molecular weight 3800) was achieved with heat-fractionation in water at 190degrees C for 5 minutes. With 0.025% NaOH, an average molecular weight of 9500 was obtained at a yield of 30%. AcGGM molecular weight standard molecules were prepared using SEC. The structure of the AcGGM was determined using H-1-NMR. The enzymatic hydrolysis of AcGGM was studied using beta-mannanase and alpha-galactosidase.

(His)6-tagged proteins were used in studies to evaluate the purification performance of HisTrap ™ HP columns. (His)6-tagged maltose-binding protein (MBP-[His]6, Mr ~43 000) expressed in E. coli was purified on HisTrap HP 1 ml columns at flow rates of 1, 2, or 4 ml/min. The yield of MBP-(His)6 was lower at the higher flow rates, but still satisfactory. Sample purity remained high and unaffected. A (His)6-tagged mannanase (Mr ~100 000) and a hydrolase (Mr ~34 000), were expressed in E. coli and P. pastoris, respectively. The (His)6-mannanase was purified using a two-step protocol with a HisTrap HP column followed by gel filtration on Superdex ™ 200 10/300 GL; (His)6-hydrolase was purified using a one-step IMAC protocol on a HisTrap HP column. High yield and purity of these (His)6-tagged enzymes were observed. The results of this study show that the performance of Ni Sepharose ™ High Performance, the prepacked medium in HisTrap HP columns, was not limited by flow rate, molecular weight of the three (His)6-tagged proteins used, or the two expression systems employed.