Unexpected observation of concentration-dependent dissociation rates for antibody-antigen complexes and other macromolecular complexes in competition experiments

J Immunol Methods. 2003 May 1;276(1-2):129-34. doi: 10.1016/s0022-1759(03)00060-7.


The dissociation rate constant of bimolecular complexes between macromolecules (k(off)) is often measured in solution by competition experiments and is generally expected to follow first-order kinetics.When measuring k(off) constants by competition for three complexes of high-affinity recombinant antibody fragments with the cognate antigen and for one calmodulin/peptide complex, a surprising dependence between apparent dissociation rate and concentration of competitor (antigen or calmodulin-binding peptide) was observed. Our results may be characteristic for macromolecules consisting of two domains (such as single-chain Fv fragments) and may reflect a transient opening of the two domains which are involved in the binding reaction, and which are connected by a polypeptide linker.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Antibodies / immunology
  • Antigen-Antibody Complex / immunology*
  • Antigens / immunology
  • Binding, Competitive
  • Calmodulin / metabolism
  • Dose-Response Relationship, Immunologic
  • Enzyme-Linked Immunosorbent Assay
  • Fibronectins / immunology
  • Kinetics
  • Ligands
  • Macromolecular Substances
  • Models, Immunological
  • Muramidase / immunology
  • Peptides / metabolism
  • Protein Binding


  • Antibodies
  • Antigen-Antibody Complex
  • Antigens
  • Calmodulin
  • Fibronectins
  • Ligands
  • Macromolecular Substances
  • Peptides
  • Muramidase