How to use immobilized metal ion affinity chromatography

Joy Winzerling, Patrick Berna, Jerker Porath

Research output: Contribution to journalArticle

50 Citations (Scopus)

Abstract

Porath and co-workers first described immobilized metal ion affinity chromatography (IMAC) in 1975 (Nature268, 598-599). Since that time IMAC has been used to isolate proteins, peptides, and nucleic acids and for cell separation. In IMAC, the metal ion is immobilized to a solid support matrix via coordinate bonding with a chelating agent. Similarly, retention of a solute on the column depends upon formation of coordinate bonds between electron-donating species of the solute in the mobile phase and the immobilized metal ion. The best-studied interaction of this nature is protein or peptide adsorption by coordinate bonding of the imidazole nitrogen of histidine side chains with certain metal ions. By judicious selection of the chelating agent, the metal ion, and the chromatography conditions, IMAC can be tailored to isolate desired constituents. To this end, we list several chelating agents that have been used in IMAC and introduce a new chelating agent, tris(2-aminoethyl)amine (Tren) and we provide protocols for activation of the matrix and the coupling of Tren. We also discuss the basic principles of IMAC to provide ground rules for initiating protein separation. Finally, we share a few applications of IMAC that demonstrate the substantive potential of this rapidly developing separation technology.

Original languageEnglish (US)
Pages (from-to)4-13
Number of pages10
JournalMethods
Volume4
Issue number1
DOIs
StatePublished - 1992

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Affinity chromatography
Ion chromatography
Affinity Chromatography
Metal ions
Metals
Ions
Chelating Agents
Peptide Nucleic Acids
Proteins
Cell Separation
Chromatography
Histidine
Adsorption
Nitrogen
Chemical activation

ASJC Scopus subject areas

  • Molecular Biology

Cite this

How to use immobilized metal ion affinity chromatography. / Winzerling, Joy; Berna, Patrick; Porath, Jerker.

In: Methods, Vol. 4, No. 1, 1992, p. 4-13.

Research output: Contribution to journalArticle

Winzerling, Joy ; Berna, Patrick ; Porath, Jerker. / How to use immobilized metal ion affinity chromatography. In: Methods. 1992 ; Vol. 4, No. 1. pp. 4-13.
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