Figure 11.

The sMF6p/FhHDM-1 protein and the peptide sFhMF6a derived from its N-terminal region are able to hemolyze RBCs preconditioned with hemin. A, hemolysis assays showing the effect of the sMF6p/FhHDM-1 protein or derived peptides and hemin on RBCs. The assay was carried out in U-shaped wells of microtiter plates and photographed after 6 h of incubation at room temperature. Wells in column 1 contain protein/peptides incubated with RBCs preconditioned with hemin, wells in column 2 contain protein/peptides preincubated with hemin and subsequently added to RBCs, wells in columns 3 and 4 are the same as in columns 1 and 2 after centrifuging at 800 × g, wells in column 5 contain protein/peptides incubated with hemin alone, and wells in column 6 contain protein/peptides incubated with RBCs alone. The proteins/peptides tested by rows were as follows: A, sMF6p/FhHDM-1; B, sFhMF6a; C, sFhMF6c. OVA (D) and LSZ (E) were used as controls. Proteins/peptides were incubated at 40 μm, hemin was incubated at 20 μm, and RBCs were incubated at 0.5% in TBS. Control wells containing RBCs plus hemin and RBCs only were placed in wells 7A and 7B, respectively. B, table depicting the degree of hemolysis of hemin-preconditioned RBCs produced by different combinations of sMF6p/FhHDM-1 protein and hemin concentrations. The values depict the degree of hemolysis, ranging from no hemolysis (−) to intermediate (++ and +++) and complete hemolysis (++++) as determined by macroscopic observation of the turbidity of RBCs (0.5%) preconditioned with 2.5–40 μm hemin and subsequently incubated with different concentrations of the sMF6p/FhHDM-1 protein (2.5–40 μm).