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Protein-Drug Binding: Determination Methods01:22

Protein-Drug Binding: Determination Methods

Determining protein-drug binding can be achieved through indirect and direct methods, each providing valuable insights into the interaction between proteins and drugs.
Indirect methods involve isolating the bound drug from its free form in biological samples such as blood, serum, or plasma. These techniques aim to measure the percentage of drugs bound to proteins. Equilibrium dialysis is a commonly used method where the free drug concentration at equilibrium is measured by separating the bound...

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Comparison of different protein concentration techniques within preformulation development.

Annette Eppler1, Markus Weigandt, Sandra Schulze

  • 1Merck Serono, CMC Development, Frankfurter Straße 250, 64293 Darmstadt, Germany.

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Small-scale antibody concentration methods were evaluated for pharmaceutical formulation. Centrifugal filtration showed the best comparability to tangential flow filtration for high-quality, stable antibody solutions.

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Area of Science:

  • Biopharmaceutical Manufacturing
  • Formulation Development
  • Protein Chemistry

Background:

  • Highly concentrated antibody solutions are crucial in the pharmaceutical industry.
  • Tangential flow filtration (TFF) is standard for production but unsuitable for small-scale early development.
  • Alternative small-scale techniques like dialysis, stirred cells, and centrifugal filters are needed.

Purpose of the Study:

  • To compare small-scale antibody concentration techniques (dialysis, stirred cells, centrifugal filters) with TFF.
  • To assess the quality and stability of highly concentrated antibody solutions produced by different methods.
  • To determine achievable protein concentrations and product characteristics.

Main Methods:

  • Model antibody solutions (10 mg/ml) were concentrated using dialysis, stirred cells, centrifugal filters, and TFF.
  • Achievable concentrations up to 100 mg/ml were assessed.
  • Stability analysis of 100 mg/ml solutions after 8-week storage (25°C and 40°C) using turbidity, SEC, SDS-PAGE, and IEF.

Main Results:

  • All techniques achieved concentrations above 100 mg/ml, with varying product qualities.
  • Dialysis resulted in the least instability.
  • Centrifugal filtration demonstrated the best comparability to TFF in IEF, turbidity, and SEC analyses.

Conclusions:

  • Small-scale techniques can yield high antibody concentrations (>100 mg/ml).
  • Centrifugal filtration is a suitable alternative to TFF for early-stage formulation development, offering comparable product quality.
  • Stability issues are primarily observed after storage, not immediately post-concentration, across all methods.