Purification of (His)6EcoRV by Metal-Chelate Affinity Chromatography (CAT#: STEM-ACM-0191-CJ)

Immobilized metal-affinity chromatography (IMAC) is a separation technique that has proven to be an efficient and versatile technology for the isolation and purification of industrial enzymes as well as proteins that are of commercial importance or used in research fields, such as genetics, molecular biology, and biochemistry. IMAC is based on the formation of immobilized metal complexes, formed by the reaction between metal ions with chelating ligands that are attached to a chromatographic support. The separation of proteins is possible through interaction of specific metal binding sites in the molecule (amino acid side chains on the protein surface or specifically developed tags, mainly histidine residues).

Biochemistry

1. Preparation of Column: The column is loaded with solid support such as sepharose, agarose, cellulose etc.. Ligand is selected according to the desired isolate. Spacer arm is attached between the ligand and solid support.
2. Loading of Sample: Solution containing a mixture of substances is poured into the elution column and allowed to run at a controlled rate.
3. Elution of Ligand-Molecule Complex: Target substance is recovered by changing conditions to favor elution of the bound molecules.

• Sample: Plants; Natural Food; Protein; Drug; Pollutants; Blood; Saliva; Serum; Plasma; Antibodies; Viruses & More
• Equipment: Agarose; Silica gel; Aluminium oxide; Acrylate; Organic polymers; Wash Buffer
• (Optional): Ligand; Spacer arm; Column

1. In the 1970s, metal-chelate affinity chromatography, also known as “immobilized-metal (ion) affinity chromatography” (IMAC), was applied for the first time.
2. Metal-chelate chromatography technology harnesses selective interactions and affinity between transition metal immobilised on a solid substrate (resin) and amino acid residues that act as electron donors in the target protein.