A comparative study of monoclonal antibodies (Mabs) purified from cell culture supernatant on EDTPA-modified zirconia beads and protein A-hyper D support

Anu Subramanian, Blanca Martinez, Jill Holm, Peter Carr, Clayton McNeff

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1 Scopus citations

Abstract

Colloidal zirconia was spray dried to yield zirconia particles, which were further modified with N, N, N′, N′- Ethylenediamine tetra methylenephosphonic acid (EDTPA) to yield a support for use in bioseparations. EDTPA modified zirconia particles will be further referred to as, r_PEZ. Cell culture supernatants rich in monoclonal antibody (Mab) subtypes IgG1, IgG2a, IgG2b, and IgG3 were chromatographed on a r_PEZ column, and on a protein A-hyper D column that was purchased commercially. All Mab subtypes bound to r_PEZ and process yields in the range of 88 to 99% were obtained. The purity of the Mab products were ascertained by gel electrophoretic analysis and were estimated to be greater than 95%. The purified Mab products obtained from r_PEZ and protein A columns were compared to the reference Mab standard in biological and enzymatic assays. The value of the dissociation constant (Kd) was found to be comparable and was in the range to that obtained with reference Mab standard (0.231 ± 0.03 M). In addition, Mabs purified with r_PEZ had the same deglycosylation profile as the reference Mab standard. Thus, it appears that the r_PEZ purified Mab is similar in activity to Mab purified with a protein A support and in addition, the zirconia surface does not adversely impact the activity of the purified Mab.

Original languageEnglish (US)
Pages (from-to)471-484
Number of pages14
JournalJournal of Liquid Chromatography and Related Technologies
Volume29
Issue number4
DOIs
StatePublished - Mar 9 2006

Keywords

  • Biological activity
  • Disassociation constant (K)
  • EPTPA
  • Monoclonal antibody
  • Protein-A
  • Zirconia

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