Isolation of Monoclonal Antibody Charge Variants by Displacement Chromatography
This unit discusses the important parameters in designing and optimizing a separation of monoclonal antibody (mAb) charge variants from process streams by ion‐exchange displacement chromatography, including sample preparation and selection of matrix, column, and appropriate buffer. A protocol is provided for determination of optimal column binding and displacement conditions, including cleaning and regeneration of the displacement columns. Curr. Protoc. Protein Sci. 69:8.10.1‐8.10.13. © 2012 by John Wiley & Sons, Inc.
Keywords: displacement chromatography; ion‐exchange chromatography; monoclonal antibodies; charge variants; immunogenicity; quality by design
Table of Contents
- Strategic Planning
- Basic Protocol: Displacement Chromatography of Monoclonal Antibody Charge Variants Using Source 15S Ion‐Exchange Resin
- Support Protocol: Analysis of Cation‐Exchange Displacement Chromatography Fractions by Second Dimension–Analytical Cation‐Exchange HPLC
- Literature Cited
Basic Protocol: Displacement Chromatography of Monoclonal Antibody Charge Variants Using Source 15S Ion‐Exchange Resin
NOTE: Longer columns tend to provide better recoveries. The aspect ratio (length to width) should be at least 20:1. Flow rates are low, so there are rarely problems with high backpressure.
Figure 8.10.1 Calculation of column binding capacity from displacement chromatogram of mAb preparative run on 4.6‐ × 250‐mm Source S‐15 column.
Figure 8.10.2 Displacement chromatogram of mAb preparation illustrating location and typical retention patterns of acidic and basic variants.
Figure 8.10.3 Displacement chromatogram of mAb prep on a 4.6‐ × 250‐mm Source 15‐S cation‐exchange column.
Figure 8.10.4 Two‐dimensional chromatogram of selected fractions from a displacement chromatography run of a mAb preparation on a 4.6‐ × 250‐mm Source 15‐S cation‐exchange column run in second dimension utilizing elution mode (same column).
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