His•Tag and GST•Tag Protein Purification Overview

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		His�Tag and GST�Tag

		GST�Tag Affinity Resins and Purification Kits
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		His�Bind Matrix Selection Guide
		His�Tag Affinity Resins and Purification Kits
		His�Tag Fusion Protein Detection Products
		His�Tag Fusion Protein Purification Products
		Protein Purification Overview

Protein Purification Overview

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Protein Purification Overview
Complete range of reagents and kits for extraction and purification

Affinity Purification
Affinity purification is based on the specific interaction of a target molecule with an immobilized ligand. Affinity technology can be used to isolate specific molecules from a mixture (e.g., His•Bind� purification), capture a desired molecule for interaction studies (e.g., immobilization on GST•Bind™ Resin), or remove a component from a reaction (e.g., protease removal with the Cleavage Capture Kits). For recombinant proteins, the addition of fusion tags using appropriate expression vectors enables affinity purification by a number of strategies. Our selection of immobilized metal affinity chromatography (IMAC) matrices for purification of His•Tag� fusion proteins is the widest available, including agarose-based His•Bind Resin; pre-charged, nonleaching Ni-NTA His•Bind Resin; magnetic based His•Mag™ Agarose Beads; cellulose-based, pre-charged His•Bind Quick Columns and Cartridges; and methacrylate His•Bind Fractogel� Resin, which enables higher pressures and capacities. Other agarose-based affinity resins enable rapid purification of GST•Tag™, S•Tag™, and T7•Tag� fusion proteins.

For the convenient, reproducible preparation of custom affinity supports containing antibodies, peptides, proteins, or other user-defined ligands, Novagen offers an activated matrix ready for coupling. PreACT™ Agarose ALD is a preparation of aldehyde-activated cross-linked agarose that forms stable amide linkages with free amino groups of proteins or peptides in the presence of sodium cyanoborohydride.

Streptavidin Agarose and MagPrep� Streptavidin Beads provide a choice of formats for purification of biotin-containing molecules. Both supports are qualified for high biotin binding activity and low non-specific background, and are well suited to a variety of applications that make use of the extraordinary affinity of biotin for streptavidin.

The MagPrep Anti-Ig Beads consist of a superparamagnetic iron oxide core encapsulated within a polystyrene shell coated with antibodies against mouse, rat, or human IgG or IgM. These beads offer many advantages for highly selective capture and immobilization of antibodies and immune complexes. Protein A and Protein G agaroses are indispensible tools for affinity purification and immunoprecipitation of antibodies and their bound antigens. Two types of supports are available that contain highly purified Protein A and/or Protein G Plus; the first is designed for purification and the other, for immunoprecipitation.

Separation Devices

For optimal performance and convenience, Novagen offers a range of devices for use with corresponding affinity resins. Three different Magnetight™ magnetic separation devices are available, designed for use with 96-well plates, multiple 1.5-ml centrifuge tubes, and 15- or 50-ml tubes. The Vacuum Manifold allows simultaneous vacuum processing of up to 12 cartridges and/or columns having Luer fittings, and is compatible with His•Bind Quick Cartridges and Columns.

Site-specific Proteases and Cleavage Capture Kits

Restriction grade site-specific proteases (Thrombin, Biotinylated Thrombin, Factor Xa, and Recombinant Enterokinase) and Cleavage Capture Kits for each enzyme offer convenient removal of fusion tags. EKapture™ and Xarrest™ Agaroses are used for the quantitative removal of rEK and Factor Xa, respectively, following cleavage of fusion proteins. Biotinylated thrombin is removed with Streptavidin Agarose.

Protein Refolding

Some purification strategies take advantage of the formation of insoluble inclusion bodies, which can sequester the target protein and protect it from degradation in the cell. To obtain the target protein in an active, soluble form, the inclusion bodies must be solubilized and the protein refolded. While the optimal conditions for solubilization and refolding largely depend on the individual protein, there are some general protocols and reagents that have been successfully used with a number of different proteins. The Protein Refolding Kit combines the reagents to perform one such protocol and can serve as a platform for using additional reagents, such as the NDSBs, for optimization.