Monovalent binding of autoantibodies to β2-glycoprotein I, detected using surface plasmon resonance at low antigen density
Véronique Regnault,
Emmanuel De Maistre,
Denis Wahl,
Thomas Lecompte,
Véronique Regnault
UMR CNRS 7563, Laboratoire d'Hématologie, Faculté de Médecine, Hémostase-Hématologie et Service de Médecine H du CHU, Nancy, France
Search for more papers by this authorEmmanuel De Maistre
UMR CNRS 7563, Laboratoire d'Hématologie, Faculté de Médecine, Hémostase-Hématologie et Service de Médecine H du CHU, Nancy, France
Search for more papers by this authorDenis Wahl
UMR CNRS 7563, Laboratoire d'Hématologie, Faculté de Médecine, Hémostase-Hématologie et Service de Médecine H du CHU, Nancy, France
Search for more papers by this authorThomas Lecompte
UMR CNRS 7563, Laboratoire d'Hématologie, Faculté de Médecine, Hémostase-Hématologie et Service de Médecine H du CHU, Nancy, France
Search for more papers by this authorVéronique Regnault,
Emmanuel De Maistre,
Denis Wahl,
Thomas Lecompte,
Véronique Regnault
UMR CNRS 7563, Laboratoire d'Hématologie, Faculté de Médecine, Hémostase-Hématologie et Service de Médecine H du CHU, Nancy, France
Search for more papers by this authorEmmanuel De Maistre
UMR CNRS 7563, Laboratoire d'Hématologie, Faculté de Médecine, Hémostase-Hématologie et Service de Médecine H du CHU, Nancy, France
Search for more papers by this authorDenis Wahl
UMR CNRS 7563, Laboratoire d'Hématologie, Faculté de Médecine, Hémostase-Hématologie et Service de Médecine H du CHU, Nancy, France
Search for more papers by this authorThomas Lecompte
UMR CNRS 7563, Laboratoire d'Hématologie, Faculté de Médecine, Hémostase-Hématologie et Service de Médecine H du CHU, Nancy, France
Search for more papers by this author Thomas Lecompte, UMR CNRS 7563, Laboratoire d'Hématologie, Faculté de Médecine, Boite Postale 184, 54505 Vandoeuvre-lès-Nancy, France. E-mail: [email protected]
Abstract
The precise mechanism of interaction between autoantibodies and β2-glycoprotein I (β2GPI) and the experimental conditions to be used for their detection are still under debate. Until now, these interactions have been studied under static conditions. We have investigated the interactions of purified IgG from 25 lupus anticoagulant-positive patients with immobilized β2GPI under flow conditions by real-time analysis based on surface plasmon resonance technology. Sensor chips were coated with purified human β2GPI coupled to dextran via amino groups at low densities (1·4, 1·8 or 2·4 ng β2GPI/mm2). Four patients' IgG displayed efficient binding and had the highest so-called antiphospholipid IgG levels by enzyme-linked immunosorbent assay (ELISA) and the highest absorbance values in an anti- β2GPI ELISA at a β2GPI density reported to be around 12 ng/mm2. Binding of antibodies to the β2GPI sensor chips proved to be dependent upon the IgG concentration and β2GPI density and was inhibited by a rabbit antibody against β2GPI. Similar association and dissociation profiles were observed for the four efficient binders. The fast rate of dissociation limited the binding of autoantibodies to β2GPI and was highly suggestive of a monovalent association, confirmed by binding of Fab fragments under similar experimental conditions. In conclusion, monovalent binding of low-affinity antibodies to β2GPI immobilized at a density as low as 1·8 ng/mm2 could be detected using surface plasmon resonance.
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