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

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

Research output: Contribution to journalArticle

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 in vitro 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 117 mg/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 60 min 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
DOIs
Publication statusPublished - 2 May 2015

Fingerprint

Cryogels
Immobilized Proteins
Staphylococcal Protein A
Adsorbents
Antigens
Antibodies
Polyacrylates
Monoclonal Antibodies
Phosphates
anthrax toxin
Ligands
Glycosylation
Exotoxins
Mechanical stability
Biomolecules
Bacilli
Fibroblasts
Cytotoxicity
Hydroxyl Radical

Bibliographical note

© 2015 The Authors. Published by Elsevier Ltd. This is an open access article under the CC BY-NC-ND license (http://creativecommons.org/licenses/by-nc-nd/4.0/).

Keywords

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

Cite this

Ingavle, Ganesh ; Baillie, Les W.J. ; Zheng, Yishan ; Lis, Elzbieta K. ; Savina, Irina ; Howell, Carol ; Mikhalovsky, Sergey ; Sandeman, Susan. / Affinity binding of antibodies to supermacroporous cryogel adsorbents with immobilized protein A for removal of anthrax toxin protective antigen. In: Biomaterials. 2015 ; Vol. 50. pp. 140-153.
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Affinity binding of antibodies to supermacroporous cryogel adsorbents with immobilized protein A for removal of anthrax toxin protective antigen. / Ingavle, Ganesh; Baillie, Les W.J.; Zheng, Yishan; Lis, Elzbieta K.; Savina, Irina; Howell, Carol; Mikhalovsky, Sergey; Sandeman, Susan.

In: Biomaterials, Vol. 50, 02.05.2015, p. 140-153.

Research output: Contribution to journalArticle

TY - JOUR

T1 - Affinity binding of antibodies to supermacroporous cryogel adsorbents with immobilized protein A for removal of anthrax toxin protective antigen

AU - Ingavle, Ganesh

AU - Baillie, Les W.J.

AU - Zheng, Yishan

AU - Lis, Elzbieta K.

AU - Savina, Irina

AU - Howell, Carol

AU - Mikhalovsky, Sergey

AU - Sandeman, Susan

N1 - © 2015 The Authors. Published by Elsevier Ltd. This is an open access article under the CC BY-NC-ND license (http://creativecommons.org/licenses/by-nc-nd/4.0/).

PY - 2015/5/2

Y1 - 2015/5/2

N2 - 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 in vitro 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 117 mg/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 60 min 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.

AB - 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 in vitro 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 117 mg/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 60 min 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.

KW - Supermacroporous cryogel

KW - Anthrax toxin protective antigen (PA)

KW - Protein A affinity cryogels

KW - Anthrax toxin specific monoclonal antibodies

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DO - 10.1016/j.biomaterials.2015.01.039

M3 - Article

VL - 50

SP - 140

EP - 153

JO - Biomaterials

JF - Biomaterials

SN - 0142-9612

ER -