Antibodies, A Laboratory Manual, Second Edition, Edited by Edward A. Greenfield



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Preparing His-Fusion Proteins from Bacteria

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Poly-histidine tags (6 or 10), unlike GST tags, are small and rarely interfere with the function, activity, or structure of target proteins. They are poorly immunogenic, unlike large tags such as GST; hence, they are widely used for the generation of fusion proteins that are used to immunize the host during antibody development. Histidine-tagged fusion proteins can be expressed in E. coli as well as in mammalian expression systems and are purified using nickel Sepharose media. Purification of His-tagged proteins is mainly based on the chelating affinity of polyhistidine for metal ions such as nickel (Ni2), cobalt, and copper. This is known as immobilized metal affinity chromatography (IMAC).

Antibodies: A Laboratory Manual, Second edition
Antibodies: A Laboratory Manual, Second edition
Antibodies: A Laboratory Manual, Second edition

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