Characterization of the interaction of the anti-properdin
IgG with their antigen by Surface plasmon resonance (SPR)
The interaction of the IgG with properdin was analyzed in real time
using a ProteOn XPR36 SPR equipment (BioRad, Marne-la-coquette, France)
and BiaCore2000 (GE Healthcare, France). Properdin was covalently
immobilized to a GLC sensor chip (BioRad) following the manufacturer’s
procedure. Alternatively, CM5 chips for BiaCore were used. Protein G
purified IgG from LN patients or healthy donors were injected for 300s
at 6 different concentrations (300, 150, 75, 35, 17.5 and 0 µg/ml)
diluted in PBS 0.005% Tween or 10mM Hepes, 145mM NaCl, 0,005% Tween 20
running buffers. The dissociation was followed for 300s. Bound protein
was regenerated with 1M NaCl, 50mM NaOH regeneration buffer.