Bivalent binding on cells varies between anti-CD20 antibodies and is dose-dependent

Sina Bondza, Toine ten Broeke, Marika University, Jeanette Leusen, Jos Buijs*

*Corresponding author for this work

Research output: Contribution to journalArticleAcademicpeer-review


Based on their mechanism of action, two types of anti-CD20 antibodies are distinguished: Type I, which efficiently mediate complement-dependent cytotoxicity, and Type II, which instead are more efficient in inducing direct cell death. Several molecular characteristics of these antibodies have been suggested to underlie these different biological functions, one of these being the manner of binding to CD20 expressed on malignant B cells. However, the exact binding model on cells is unclear. In this study, the binding mechanism of the Type I therapeutic antibodies rituximab (RTX) and ofatumumab (OFA) and the Type II antibody obinutuzumab (OBI) were established by real-time interaction analysis on live cells. It was found that the degree of bivalent stabilization differed for the antibodies: OFA was stabilized the most, followed by RTX and then OBI, which had the least amount of bivalent stabilization. Bivalency inversely correlated with binding dynamics for the antibodies, with OBI displaying the most dynamic binding pattern, followed by RTX and OFA. For RTX and OBI, bivalency and binding dynamics were concentration dependent; at higher concentrations the interactions were more dynamic, whereas the percentage of antibodies that bound bivalent was less, resulting in concentration-dependent apparent affinities. This was barely noticeable for OFA, as almost all molecules bound bivalently at the tested concentrations. We conclude that the degree of bivalent binding positively correlates with the complement recruiting capacity of the investigated CD20 antibodies.
Original languageEnglish
Article number1792673
Pages (from-to)1-13
Number of pages13
Issue number1
Publication statusPublished - 4 Aug 2020


  • Affinity
  • CD20
  • binding kinetics
  • cell-based assay
  • obinutuzumab
  • ofatumumab
  • receptor-ligand interactions
  • rituximab
  • therapeutic antibodies


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