Protein A affinity precipitation of human immunoglobulin G

Lars Janoschek, Matthias Freiherr von Roman, Sonja Berensmeier

Research output: Contribution to journalArticlepeer-review

7 Scopus citations

Abstract

The potential of protein A affinity precipitation as an alternative method for traditional antibody purification techniques was investigated. Recombinant produced protein A from Staphylococcus aureus (SpA) was covalently linked to the pH-responsive copolymer Eudragit® S-100 and used for purification of human immunoglobulin G (hIgG). The Eudragit-SpA conjugate had a static binding capacity of 93.9±2.8mg hIgG per g conjugate and a dissociation constant of 787±67nM at 7±1°C. The antibody was adsorbed rapidly onto Eudragit-SpA and reached equilibrium within 5min. An excess of hIgG binding sites, provided by the conjugate, as well as adjusted elution conditions resulted in an appropriate hIgG purification performance. In summary, Eudragit-SpA was successfully applied to capture hIgG from a protein mixture with 65% antibody yield in the elution step. Nearly 96% purity and a purification factor of 12.4 were achieved. The Eudragit-SpA conjugate showed a stable ligand density over several cycles, which enabled reusability for repeated precipitation of hIgG. According to this, pH induced affinity precipitation can be seen as a potential alternative for protein A chromatography in antibody purification processes.

Original languageEnglish
Pages (from-to)72-78
Number of pages7
JournalJournal of Chromatography B: Analytical Technologies in the Biomedical and Life Sciences
Volume965
DOIs
StatePublished - 15 Aug 2014

Keywords

  • Affinity precipitation
  • Antibody purification
  • Eudragit
  • Immunoglobulin G
  • Protein A

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