Fig. 4. Merozoite-specific IgM inhibits invasion in a complement-dependent manner.

(A) Isolated IgM and IgG fractions at 50 μg/ml from malaria-exposed donors (Sabah, Malaysian Borneo) or naïve donors (Australia) were tested in merozoite invasion inhibition assays (at 50 μg/ml) with normal serum (NS) or heat-inactivated serum (HIS) from malaria-naïve donors as a source of complement. Data are expressed as the percentage of invasion in heat-inactivated serum, with no antibody control. Data are means ± SD of two assays performed in duplicate. Significant differences are indicated as P < 0.05 by Mann-Whitney U test. *P < 0.05. (B) Isolated IgM and IgG fractions were tested for deposition of complement components (C1q, C3b, and C5b-C9) on intact merozoites or recombinant MSP2 by ELISA with 10% normal sera from malaria-naïve donors as a source of complement. Data represent the level of complement binding measured in OD values minus deposition observed using naïve IgM and IgG. (C) IgM and IgG fractions were tested for deposition of complement components on recombinant MSP2 by ELISA with 10% C1q-depleted sera and C1q-depleted sera reconstituted with purified C1q (10 μg/ml). Data are means ± SEM of two independent assays performed in duplicate. (D) The level of C1q-fixing antibodies and IgM and IgG1/IgG3 isotypes to merozoites was quantified in 50 individuals with a clinical malaria episode from Sabah. IgM, IgG1, and IgG3 were correlated with C1q fixation. (E) To compare directly between C1q fixation and IgM or IgG1/IgG3, antibody levels were converted to arbitrary units (the percentage of the highest measured response within cohort) and ranked according to levels of C1q fixation. In some individuals with relatively high C1q-fixing antibodies (defined as >40% of highest responder), IgM dominated the antibody response targeting the merozoite.