Figure 1. CC activate the alternative- and classical complement pathways.

Human serum was incubated at 37°C, for indicated times, in the presence of CC (3 × 10⁷ particles/ml), zymosan and heat aggregated IgG (Zym-IgG) or PBS. (A) The end product in complement activation, TCC, showed a significant increase (***, p < 0.001) in response to CC compared to PBS at every time points, (B) the activation product C3bc, from the common complement component C3 for all three initial pathways, showed a significant increase (***, p < 0.001) at all time points in response to CC compared to PBS, (C) the alternative pathway convertase, C3bBbP, showed a significant increase (***, p < 0.001) in response to CC compared to PBS at every time points. The increase in (D) the common activation product for the classical and lectin pathways, C4bc, and (E) the activation product for the classical pathway, C1rs-C1-INH, did not reach statistical significance. Data plotted are mean ± SEM from six independent experiments with serum from healthy donors. (F) C1q binding to CC, when incubated in plasma for 30 minutes, measured by flow cytometry. (G) TCC in C1q depleted serum (C1q dep.) with or without reconstitution (C1q rec.) (*, p < 0.05) with purified C1q upon CC (1.5 × 10⁷ particles/ml) stimulation, measured by ELISA. One out of six independently performed experiments is shown. In the lower panel, human whole blood was incubated with CC (1.5 × 10⁷ particles/ml) for 30 minutes. (H) Binding of TCC to the crystals was detected using anti C5b-9 and anti-mouse IgG conjugated with Alexa-488, and for C3bc (I) anti-C3bc antibody directly conjugated to FITC. (J) Control IgG2a conjugated to FITC. Scale bars represent 10 µm. Data are representative of two independent experiments. AU = arbitrary units. DIC = Differential Interference Contrast.