Purification of human plasma fibronectin using immobilized gelatin and Arg affinity chromatography

Pietro Speziale, Livia Visai, Simonetta Rindi, Antonella Di Poto

Research output: Contribution to journalArticlepeer-review

Abstract

This protocol describes a method for purification of fibronectin (Fn) from human plasma based on a combination of gel filtration and affinity chromatography steps. Clarified plasma is first loaded onto a Sepharose CL-4B column and unbound material is sequentially purified on columns containing covalently coupled gelatin and Arg. The elution conditions are optimized to obtain a homogeneous preparation of Fn on sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). Although the Fn yield is expected to be lower than that obtained using other methods, affinity adsorbents based on gelatin and Arg and gentle elution steps offer advantages including a high purity of the preparation and a correctly folded protein. The preparation can be useful for interaction studies and analysis of biological and immunological activities of Fn.

Original languageEnglish
Pages (from-to)525-533
Number of pages9
JournalNature Protocols
Volume3
Issue number3
DOIs
Publication statusPublished - Mar 2008

ASJC Scopus subject areas

  • Biochemistry, Genetics and Molecular Biology(all)

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