Fig. 2.

Complement opsonisation is required for monocyte phagocytosis of antibody-opsonised IE a Monocyte phagocytosis of CS2-IE (Unop) or CS2-IE opsonised with rabbit anti-human RBC antibody (OP) was determined using whole blood, whole blood reconstituted to its original volume with autologous heat-inactivated plasma (HI plasma) or with autologous plasma (plasma) as indicated. Phagocytosis by classical CD14^hiCD16- (left hand panel), intermediate CD14^hiCD16+ (middle panel) and non-classical CD14^loCD16+ (right hand panel) monocytes were measured. Data represent mean (sem) of independent experiments using blood from three separate donors. Differences between conditions were assessed by one-way ANOVA using Tukey’s test for multiple comparisons. b Unopsonised CS2-IE or CS2-IE opsonised with rabbit anti-human RBC antibody were added to heparinised plasma for 0 and 30 minutes at 37 °C, stained with antiC3, and RBC analysed by flow cytometry. Histograms represent C3 staining at 0 time (dark grey histogram) or 30 minutes (light grey histogram). Right hand panel represents C3 staining of CS2-IE opsonised with rabbit anti-human RBC antibody after incubation in heparinised plasma for the indicated times at 4 °C (solid black circles) or 37 °C (open circles). c Compstatin (R&D Systems) was added to whole blood from stock solutions dissolved in PBS at the indicated final concentrations then phagocytosis of CS2-IE opsonised with rabbit anti-human RBC antibody by intermediate (open circles) or classical monocytes (solid black circles) determined (left hand panel) or non-classical monocytes (open squares) determined (right hand panel). The absolute values of phagocytosis by non-classical monocytes were very low and hence these data are plotted separately. Data represent mean (sem) of independent experiments using blood from three separate donors. ANOVA analysis of variance, IE infected erythrocytes, RBC red blood cells, sem standard error of the mean