A gel-based method for purification of apolipoprotein A-I from small volumes of plasma

Rachel J. Brace, Brie Sorrenson, Dmitri Sviridov, Sally P.A. McCormick

Research output: Contribution to journalArticleResearchpeer-review

15 Citations (Scopus)

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.

Original languageEnglish
Pages (from-to)3370-3376
Number of pages7
JournalJournal of Lipid Research
Volume51
Issue number11
DOIs
Publication statusPublished - Nov 2010
Externally publishedYes

Keywords

  • Cholesterol efflux
  • Gel purification
  • High density lipoproteins
  • Size-exclusion chromatography

Cite this