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A method for improving protein A chromatography's aggregate removal capability.

Yuan Zhang1, Ying Wang1, Yifeng Li1

  • 1Technology and Process Development (TPD), WuXi Biologics, 288 Fute Zhong Road, Waigaoqiao Free Trade Zone, Shanghai, 200131, China.

Protein Expression and Purification
|March 5, 2019
PubMed
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A new method enhances Protein A chromatography's ability to remove antibody aggregates. Combining salts like calcium chloride or sodium chloride with polyethylene glycol (PEG) significantly improves monomer-aggregate separation during purification.

Area of Science:

  • Biopharmaceutical purification
  • Chromatographic techniques
  • Protein aggregate removal

Background:

  • Protein A chromatography is a primary method for antibody purification.
  • However, it exhibits limited efficacy in removing antibody aggregates under standard conditions.
  • Antibody aggregates can negatively impact therapeutic efficacy and safety.

Purpose of the Study:

  • To develop an improved method for removing antibody aggregates during Protein A chromatography.
  • To enhance the resolution between antibody monomers and aggregates.
  • To increase the overall robustness of the downstream purification process.

Main Methods:

  • Supplementing Protein A chromatography buffers (wash and elution) with specific salt/polyethylene glycol (PEG) combinations.
Keywords:
Antibody aggregate removalCalcium chlorideMonomer-aggregate resolutionPolyethylene glycol (PEG)Protein A chromatographySodium chloride

Related Experiment Videos

  • Investigating the individual and combined effects of calcium chloride/PEG and sodium chloride/PEG on aggregate removal.
  • Evaluating the synergistic impact of salt and PEG on chromatographic resolution.

    • Main Results:

    • The addition of salt (calcium chloride or sodium chloride) alone showed moderate aggregate resolution enhancement.
    • Polyethylene glycol (PEG) alone did not impact resolution.
    • The combination of salt and PEG demonstrated a significant synergistic effect, drastically improving monomer-aggregate separation.
    • Optimized conditions reduced aggregate levels in the elution pool from 20% to 3-4% compared to a control.

    Conclusions:

    • A novel method using salt/PEG combinations effectively enhances Protein A chromatography for aggregate removal.
    • This synergistic approach significantly improves the separation of antibody monomers from aggregates.
    • The developed method offers a robust solution for mitigating aggregate contamination at the critical capture step in downstream processing.