Preparations of PEMSs, intact DVs, and free hemozoin. (A) Merozoites and DVs encased in the parasitophorous vacuole from pRBCs after rupture in the presence of the protease inhibitor E64, which inhibits lysis of the parasitophorous vacuole membrane but not lysis of the erythrocyte membrane. (B) DVs isolated from trophozoites. (C) Hemozoin crystals isolated from sonicated DVs by centrifugation in Percoll. Scale bar indicates 5 μm. (D) Complement consumption tests were performed with materials in panels A through C using 10% NHS. DVs were used as a positive control and similar concentrations of heme were used throughout. Neither PEMSs nor isolated hemozoin had complement-activating capacity. One representative assay of 3 independent experiments is shown, with SD from triplicate determinations.