Effect of anti-properdin IgG on formation of C3 convertase
The effect of total IgG, positive for anti-properdin autoantibodies on the interaction of properdin with C3b, C3bB and C3bBb was analyzed in real time using a ProteOn XPR36 SPR equipment (BioRad, Marne-la-coquette, France). Properdin was covalently immobilized to a GCL sensor chip (BioRad), following the manufacturer’s procedure. Protein G purified IgG from LN patients, positive for anti-properdin autoantibodies (but negative for anti-C3b and anti-FB), and healthy donors were injected for 300s at dilution 1:15 in 10 mM Hepes, 50 mM NaCl, 10 mM MgCl, 0.005% Tween 20, pH 7.4 buffer. The same buffer was separately injected to serve as a control. Following 300s of dissociation, C3b (13µg/ml), C3b (13µg/ml) + Factor B (10µg/ml) and C3b (13µg/ml) + Factor B (10µg/ml) + Factor D (0,5µg/ml) (Complement Technology) were injected for 300s of association, followed by 300s of dissociation.