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Activated Cross-linked Agarose for the Rapid Development of Affinity Chromatography Resins - Antibody Capture as a Case Study
Published on: August 16, 2019
Two-dimensional fluorescence difference gel electrophoresis for comparison of affinity and non-affinity based
Julita K Grzeskowiak1, Anne Tscheliessnig, Poh Choo Toh
1Department of Biotechnology, University of Natural Resources and Applied Life Sciences, Vienna, Muthgasse 18, A-1190 Vienna, Austria.
Abstract:
Although Staphylococcus Protein A (SpA) affinity chromatography is the state of the art capture step for antibody purification, non-affinity methods are more economical. We used two-dimensional fluorescence difference gel electrophoresis (2-D DIGE) to evaluate the purification of a recombinant IgG(1) antibody from cultured cells, with two different processes: (1) SpA capture followed by cation-exchange chromatography (CEX); and (2) CEX capture, followed by anion exchanger, then hydrophobic interaction chromatography. Efficiencies were similar in sodium dodecylsulphate polyacrylamide gel electrophoresis and size-exclusion chromatography; however, 2-D DIGE revealed higher efficiency with SpA than with CEX capture. Thus, 2-D DIGE is a valuable tool for downstream process development.
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