Assessment of ADCC activity of therapeutic antibodies: from FcγRIIIa binding kinetics to functional cell-based assays

By
  • Fabian Vandermeers
  • Paula Paci
  • Thomas Henri
  • Arnaud Delobel
Date
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Antibody-dependent cellular cytotoxicity (ADCC) is a key effector mechanism driving the activity of many therapeutic monoclonal antibodies. It relies on the interaction between antibody-coated target cells and immune effector cells via the FcγRIIIa receptor, ultimately leading to target cell elimination. Given its complexity and dependence on multiple molecular and cellular factors, a comprehensive evaluation of ADCC requires robust and complementary analytical approaches.

This application note presents an integrated workflow combining biophysical binding assays and functional cell-based assays to characterise ADCC activity from receptor engagement through to cytotoxic response.

Explore the complete ADCC evaluation workflow in our application note below

Core analytical insights

Integrated workflow combining SPR, BLI, Lumit immunoassay, reporter assay and cell killing assay:

  • Quantitative determination of FcγRIIIa binding kinetics (ka, kd) and affinity (KD) using SPR and BLI
  • Detection of Fc–receptor interaction differences via Lumit competition assay (EC50 comparison)
  • Functional potency assessment through reporter gene activation and direct cytotoxicity assays
  • Orthogonal evaluation enabling correlation between molecular binding and biological activity

HOW THIS SUPPORTS YOUR ADCC ANALYTICAL STRATEGY?

A multi-layered understanding of ADCC is essential to ensure consistent performance and regulatory compliance of therapeutic antibodies. This integrated approach provides both mechanistic insight and functional confirmation across development stages.

  • Enables detailed characterisation of FcγRIIIa binding, including polymorphism impact (V158/F158)
  • Supports candidate selection and Fc engineering evaluation through precise affinity and kinetic data
  • Provides robust and reproducible potency assessment suitable for batch release and stability studies
  • Facilitates comparability and biosimilarity assessments with orthogonal analytical evidence
  • Confirms mechanism of action through biologically relevant cell killing assays
  • Allows efficient screening and rapid confirmation using high-throughput binding assays

Together, these techniques offer a comprehensive and scalable strategy to assess ADCC activity from early development to routine quality control.

APPLICATIONS

  • Candidate screening and selection
  • Fc engineering and glycoform comparison
  • Comparability and biosimilarity studies
  • Batch release and stability testing
  • Mechanistic investigations of ADCC activity

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