The stepwise generation of alternative pathway convertases monitored by plasmon resonance. C3b was conjugated to a CM 5 chip by amide bonds and used to create an initial alternative pathway convertase by sequential addition of factors B and D (as indicated by arrows) in the presence of 1 mmol/L Ni²⁺. The active convertase was thereafter subjected to native C3, which subsequently bound to the surface in the form of C3b. When the cycle with factors B and D was repeated, additional alternative pathway convertase complexes were generated. These convertase complexes were preincubated with Compstatin (70 μmol/L) followed by native C3 added in the presence of either Compstatin (70 μmol/L) or control peptide (70 μmol/L).