Cation exchange chromatography in antibody purification: pH screening for optimised binding and HCP removal
| Autor: | Andre Kiesewetter, Andreas Stein |
|---|---|
| Rok vydání: | 2006 |
| Předmět: |
Titration curve
medicine.drug_class Clinical Biochemistry Ion chromatography Enzyme-Linked Immunosorbent Assay Monoclonal antibody Biochemistry Analytical Chemistry Cations Cell Line Tumor medicine Animals Cation Exchange Resins Polyacrylamide gel electrophoresis Chromatography High Pressure Liquid Chromatography Ion exchange biology Chemistry Antibodies Monoclonal Reproducibility of Results Cell Biology General Medicine Hydrogen-Ion Concentration Chromatography Ion Exchange Electrophoresis biology.protein Electrophoresis Polyacrylamide Gel Leaching (metallurgy) Protein A Protein Binding |
| Zdroj: | Journal of chromatography. B, Analytical technologies in the biomedical and life sciences. 848(1) |
| ISSN: | 1570-0232 |
| Popis: | The production of pharmaceutical antibodies requires reliable and rapid processes with high purity and yield. Although protein A gels selectively and efficiently bind antibodies in the capture step, intense research is going on to find alternatives that can abolish the drawbacks of protein A chromatography. Ion exchangers e.g. are more robust, considerably cheaper and can eliminate ligand leaching. For the strong cation exchangers Fractogel EMD SO3- (M) and Fractogel EMD SE Hicap (M) we have evaluated the influence of pH for optimised binding and removal of host cell protein (HCP). In a fast initial screening we measured batch binding capacities. Subsequent scale-down to 96-well plate format proved that assay miniaturisation still provided reliable data. We demonstrated with the principle of residence time that scout columns are suitable for dynamic studies. The optimum pH range from batch binding was transferred to scout columns which were then used to screen for maximum dynamic capacities. In addition IEF titration curve analysis was employed to define a final operational pH. With this pH we ran labscale columns to purify monoclonal antibody. The cation exchangers showed high step yields and host cell proteins in the pools from gradient elution were reduced very effectively. |
| Databáze: | OpenAIRE |
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