Member since: 2015
Organization: Oklahoma State University
Great results, can't survive with this product.
Application Area: Purification of His-tagged recombinant protein after heterologous expression
"Once the resin to total protein (after cell lysis) is optimized, all of our target protein did bind to the resin very effectively. After washing, elution was also very effective providing high yield."
Thermo Scientific HisPur Ni-NTA Superflow Agarose is a nitrilotriacetic acid (NTA) modified Superflow 6 support charged with divalent nickel (Ni+2) designed for FPLC purification of poly-histidine-tagged proteins.
Features of HisPur Ni-NTA Superflow Agarose:
• High capacity—binds greater than 20 mg of His-tagged GFP per mL of resin at a linear flow rate of 300 cm/hour
• High purity—provides greater than 80% purity of eluted fractions when used to purify from lysates
• Versatile—can be used to purify proteins under native and denaturing conditions
• Robust—highly crosslinked beads tolerate linear flow rates up to 1200 cm/hour
• Compatible—maintains function in a wide variety of chemicals and pH conditions
• Cost effective—competitively priced resin is stable through multiple cycles of cleaning and reuse
HisPur Ni-NTA Superflow Agarose is a highly crosslinked, durable resin that does not compress at flow rates used for medium- or large-scale FPLC purifications. The resin holds up well to a variety of chemicals and pH values, and is compatible with common clean-in-place procedures. Compared to cobalt and other ligands used for immobilized metal affinity chromatography (IMAC), nickel provides greater capacity for His-tagged protein purification. HisPur Ni-NTA Superflow Agarose exhibits a high dynamic binding capacity across a range of flow rates, making it an excellent choice for larger scale purifications.
• Medium- to large-scale FPLC purification of polyhistidine tagged proteins
Affinity chromatography is often used as a quick and easy approach for the purification of recombinant proteins. One such method of purification involves the binding of a poly-histidine tag to a divalent metal cation that has been coordinated by a chelator immobilized on a beaded support. For immobilized metal affinity chromatography (IMAC) purification of His-tagged proteins, the type of bead, chelator, and metal immobilized influences purity, yield, and flow rate performance of these purifications.
HisPur Ni-NTA Superflow Agarose is a nitrilotriacetic acid (NTA) modified Superflow 6 support charged with divalent nickel (Ni+2) designed for FPLC purification of poly-histidine-tagged protein. To demonstrate performance, 6xHis-GFP was over expressed in a 100L reactor and the cell mass was collected in multiple fractions. A portion of the biomass (240g) was lysed with 2L of lysis buffer supplemented with Thermo Scientific Halt Protease Inhibitor (Part No. 78439). The lysate was clarified and split into two fractions. The target protein was then purified using HisPur Ni-NTA Superflow Agarose and a leading competitor's resin in equivalent 200 mL packed bead columns. Total yield, recovery, and purity (>80%) were nearly identical for both resins, and more than 4 grams of target protein was purified in less than 3 hours.