Today, most lab-scale purifications are performed with affinity-tagged proteins, such as the purification of histidine-tagged proteins using IMAC, also known as metal chelate affinity chromatography. In this technique, the medium is first charged with a divalent metal ion (e.g., Ni²⁺, Cu²⁺ , Zn²⁺, or CO²⁺), which is immobilized to the medium via a chelating ligand.

When sample is applied, proteins containing histidine, cysteine, and tryptophan residues bind the immobilized metal ions. Bound protein can be eluted by competitive elution with imidazole, for example, or by lowering the pH.

Since many proteins have intrinsic histidine and/or cysteine amino acid residues, some nonspecific binding to the IMAC media can occur. Therefore, it is often necessary to optimize binding, wash, and elution conditions by varying the concentration of imidazole in these solutions. Increasing the concentration of imidazole in the binding and wash buffers generally decreases nonspecific binding, whereas lower concentrations give stronger affinity interaction. The key is finding the right balance.

Find out more about our precharged and uncharged IMAC Media. Choose from bulk media, prefilled 96-well filter plates, and several prepacked column formats including HiTrap columns and spin columns.