title: A gel-based method for purification of apolipoprotein A-I from small volumes of plasma.
abstract: We present here a gel-based method for rapid purification of apolipoprotein A-I (apoA-I) from small volumes of human plasma. After isolation of high density lipoprotein from plasma, the apoA-I protein was separated by electrophoresis and the apoA-I band excised from the gel. The apoA-I was then eluted from the gel strip, concentrated, and delipidated ready for use. The structure and function of the gel-purified apoA-I protein was compared against apoA-I purified by the traditional size-exclusion chromatography method. The α-helical content of the gel-purified apoA-I as determined by circular dichroism was similar to chromatography-purified apoA-I. The functional activity of gel-purified apoA-I, as determined by cholesterol efflux assays in primary human fibroblasts and RAW264.7 macrophages, was also comparable with chromatography-purified apoA-I. This method is a valid alternative for apoA-I purification with some advantages over traditional chromatography purification including a much reduced plasma volume requirement, less time and cost, and a higher percentage protein recovery. The method is particularly suitable for applications requiring the purification of apoA-I from multiple human or animal samples of interest.
authors.affiliation: Department of Biochemistry, University of Otago, Dunedin, New Zealand.
authors.forename: Rachel J
authors.initials: RJ
authors.lastname: Brace
authors.forename: Brie
authors.initials: B
authors.lastname: Sorrenson
authors.forename: Dmitri
authors.initials: D
authors.lastname: Sviridov
authors.forename: Sally P A
authors.initials: SP
authors.lastname: McCormick
mesh_terms: D016632:Apolipoprotein A-I / Q000097:blood* / Q000302:isolation & purification*; D005591:Chemical Fractionation / Q000379:methods*; D005782:Gels; D006801:Humans; D007202:Indicators and Reagents / Q000737:chemistry; D007753:Laboratories
chemical_list: D016632:Apolipoprotein A-I; D005782:Gels; D007202:Indicators and Reagents