Poly-histidine tagging is widely employed for the purification of recombinant target proteins via immobilized metal affinity chromatography (IMAC). The advantages include small tag size and high affinity and specificity of poly-His tag binding to divalent metals at neutral pH. The primary disadvantage is that microbial hosts naturally express metal binding proteins that may co-purify with the target protein. Metal affinity matrices are most commonly in the form of an agarose bead, magnetic bead or a prepacked spin column. The metal ions, typically Nickel or Cobalt, are presented by a ligand with specific properties for divalent ion coordination. The nitrilotriacetic acid (NTA) ligand is most commonly employed for metal immobilization but other ligands are being adopted which offer greater stability against EDTA/metal ion leaching and greater resistance to reducing agents such as beta-mercaptoethanol and DTT.
- Protein Expression Using NiCo21(DE3) (C2529)
- Protocol for Removal of IMAC Contaminating Proteins (C2529)
- High Efficiency Transformation Protocol (C2529)
- NEBExpress® Ni Spin Columns Quick Start Protocol (NEB #S1427)
- NEBExpress® Ni Resin Quick Start Protocol (NEB #S1428)
- Quick Start Protocol for NEBExpress® Ni-NTA Magnetic Beads
- Protein Expression with T7 Express strains
- 5 Minute Transformation Protocol (C2529)
Avoid Common Obstacles in Protein Expression
Read how to avoid common obstacles in protein expression that prevent interactions with cellular machinery.
- Competent Cell Brochure
- Protein Expression & Purification Brochure
- Purification Beads, Columns and Resins Brochure
- Competent Cell Product Comparison
- Purification Beads, Columns and Resins
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