Affinity binding of antibodies to supermacroporous cryogel adsorbents with immobilized protein A for removal of anthrax toxin protective antigen

Ganesh C. Ingavle, Les W.J. Baillie, Yishan Zheng, Elzbieta K. Lis, Irina N. Savina, Carol A. Howell, Sergey V. Mikhalovsky, Susan R. Sandeman

Research output: Contribution to journalArticlepeer-review

29 Citations (Scopus)

Abstract

Polymeric cryogels are efficient carriers for the immobilization of biomolecules because of their unique macroporous structure, permeability, mechanical stability and different surface chemical functionalities. The aim of the study was to demonstrate the potential use of macroporous monolithic cryogels for biotoxin removal using anthrax toxin protective antigen (PA), the central cell-binding component of the anthrax exotoxins, and covalent immobilization of monoclonal antibodies. The affinity ligand (protein A) was chemically coupled to the reactive hydroxyl and epoxy-derivatized monolithic cryogels and the binding efficiencies of protein A, monoclonal antibodies to the cryogel column were determined. Our results show differences in the binding capacity of protein A as well as monoclonal antibodies to the cryogel adsorbents caused by ligand concentrations, physical properties and morphology of surface matrices. The cytotoxicity potential of the cryogels was determined by an invitro viability assay using V79 lung fibroblast as a model cell and the results reveal that the cryogels are non-cytotoxic. Finally, the adsorptive capacities of PA from phosphate buffered saline (PBS) were evaluated towards a non-glycosylated, plant-derived human monoclonal antibody (PANG) and a glycosylated human monoclonal antibody (Valortim®), both of which were covalently attached via protein A immobilization. Optimal binding capacities of 108 and 117mg/g of antibody to the adsorbent were observed for PANG attached poly(acrylamide-allyl glycidyl ether) [poly(AAm-AGE)] and Valortim® attached poly(AAm-AGE) cryogels, respectively, This indicated that glycosylation status of Valortim® antibody could significantly increase (8%) its binding capacity relative to the PANG antibody on poly(AAm-AGE)-protien-A column (p<0.05). The amounts of PA which remained in the solution after passing PA spiked PBS through PANG or Valortim bound poly(AAm-AGE) cryogel were significantly (p<0.05) decreased relative to the amount of PA remained in the solution after passing through unmodified as well as protein A modified poly(AAm-AGE) cryogel columns, indicates efficient PA removal from spiked PBS over 60min of circulation. The high adsorption capacity towards anthrax toxin PA of the cryogel adsorbents indicated potential application of these materials for treatment of Bacillus anthracis infection.

Original languageEnglish
Pages (from-to)140-153
Number of pages14
JournalBiomaterials
Volume50
Issue number1
DOIs
Publication statusPublished - Jan 1 2015

Keywords

  • Anthrax toxin protective antigen (PA)
  • Anthrax toxin specific monoclonal antibodies
  • Protein A affinity cryogels
  • Supermacroporous cryogel

ASJC Scopus subject areas

  • Bioengineering
  • Ceramics and Composites
  • Biophysics
  • Biomaterials
  • Mechanics of Materials

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