title: Ion-Exchange Chromatography: Basic Principles and Application.
abstract: Ion-Exchange Chromatography (IEC) allows for the separation of ionizable molecules on the basis of differences in charge properties. Its large sample-handling capacity, broad applicability (particularly to proteins and enzymes), moderate cost, powerful resolving ability, and ease of scale-up and automation have led to it becoming one of the most versatile and widely used of all liquid chromatography (LC) techniques. In this chapter, we review the basic principles of IEC, as well as the broader criteria for selecting IEC conditions. By way of further illustration, we outline basic laboratory protocols to partially purify a soluble serine peptidase from bovine whole brain tissue, covering crude tissue extract preparation through to partial purification of the target enzyme using anion-exchange chromatography. Protocols for assaying total protein and enzyme activity in both pre- and post-IEC fractions are also described.
authors.affiliation: School of Biotechnology, Dublin City University, Glasnevin, Dublin 9, Ireland. phil.cummins@dcu.ie.
authors.forename: Philip M
authors.initials: PM
authors.lastname: Cummins
authors.affiliation: School of Biotechnology, Dublin City University, Glasnevin, Dublin 9, Ireland.
authors.forename: Keith D
authors.initials: KD
authors.lastname: Rochfort
authors.affiliation: School of Biotechnology, Dublin City University, Glasnevin, Dublin 9, Ireland.|National Centre for Sensor Research, Dublin City University, Glasnevin, Dublin 9, Ireland.|Irish Separation Sciences Cluster (ISSC), Dublin City University, Glasnevin, Dublin 9, Ireland.
authors.forename: Brendan F
authors.initials: BF
authors.lastname: O'Connor
mesh_terms: D000818:Animals; D001921:Brain / Q000378:metabolism; D002417:Cattle; D011232:Chemical Precipitation; D002852:Chromatography, Ion Exchange / Q000379:methods*; D014020:Tissue Extracts / Q000737:chemistry / Q000302:isolation & purification
chemical_list: D014020:Tissue Extracts
keywords: Anion exchange; Capture; Cation exchange; Continuous-bed; DEAE-Sepharose® Fast Flow; Liquid chromatography; Polish; Prolyl oligopeptidase