Construction, purification, and characterization of anti-BAFF scFv-Fc fusion antibody expressed in CHO/dhfr- cells

Appl Biochem Biotechnol. 2009 Jun;157(3):562-74. doi: 10.1007/s12010-008-8434-6. Epub 2008 Dec 20.

Abstract

Elevated levels of B-cell-activating factor of the tumor necrosis factor family (BAFF) have been implicated in the pathogenesis of autoimmune diseases in human. In this study, we have constructed a vector for the expression of a novel compact antibody composed of anti-BAFF single-chain antibody fragment (scFv) and the Fc region (the hinge region, CH2, and CH3 domains) of human IgG1 in Chinese hamster ovary cells. The scFv-Fc fusion protein, showing spontaneous Fc fragment-mediated homodimerization via disulfide bridges, was affinity-purified on protein A Sepharose from culture supernatant. The scFv-Fc antibody was demonstrated to retain high binding affinity to antigen and prolonged clearance time in blood and to possess some human IgG crystallizable fragment effector functions such as protein A binding and antibody-dependent cellular cytotoxicity. These results suggest that this recombinant antibody may have therapeutic applications in the therapy of autoimmune disorders mediated by BAFF.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • B-Cell Activating Factor / immunology*
  • B-Cell Activating Factor / metabolism
  • Blotting, Western
  • CHO Cells
  • Cricetinae
  • Cricetulus
  • Enzyme-Linked Immunosorbent Assay
  • Humans
  • Immunoglobulin Fc Fragments / immunology*
  • Immunoglobulin Fc Fragments / metabolism
  • Immunoglobulin Fragments / immunology*
  • Immunoglobulin Fragments / metabolism
  • Models, Biological
  • Recombinant Fusion Proteins / immunology*
  • Recombinant Fusion Proteins / metabolism

Substances

  • B-Cell Activating Factor
  • Immunoglobulin Fc Fragments
  • Immunoglobulin Fragments
  • Recombinant Fusion Proteins