Subcellular distribution of PfEMMA1 in merozoites and infected RBCs. (A–H) Merozoites (Pf3D7) that were live, non-permeabilized (A–D), or permeabilized (E–H) were probed with antibodies against PfEMMA1 fragment 2 (A–C and E–G), and labeling of PfEMMA1 is shown. No proteins were detected with preimmune serum (D and H). A merozoite is in contact with an uninfected RBC at its apex where PfEMMA1 is highly expressed (E). (I–L) A live, non-permeabilized Pf3D7 schizont (I) and merozoite (J) were compared with a permeabilized schizont (K) and merozoite (L); cells were probed with anti-PfEMMA1 C-terminus antibodies (I–L). Staining of PfEMMA1 was observed only in permeabilized iRBCs (K) and merozoites (L). Counterstains included DAPI, PfAMA1, PfMSP1 and →4, PfRH5, and anti-GPC (glycophorin C). DIC, differential interference contrast microscopy. Scale bars, 2 μm. We identified the protein on the exofacial surface of non-permeabilized merozoites in close association with PfRH5 (rhoptry protein), but remote from PfAMA1 (microneme protein) and PfMSP1 (surface protein; Pearson’s correlation coefficient <0.5 for each; A–C). We were unable to detect

any proteins using preimmune sera (D). PfEMMA1 was visualized in permeabilized merozoites (E–G) that was not detected with preimmune sera (H). In certain fields (C and E), PfEMMA1 appears to be localized to the apex of merozoites and is oriented in close proximity to the site of contact with an uninfected RBC (E). Antibodies to PfEMMA1 fragment 1 showed similar staining patterns (E and F).

Michelow IC, Park S, Tsai SW, Rayta B, Pasaje CFA, Nelson S, Early AM, Frosch AP, Ayodo G, Raj DK, Nixon CE, Nixon CP, Pond-Tor S, Friedman JF, Fried M, Duffy PE, Le Roch KG, Niles JC, Kurtis JD. A newly characterized malaria antigen on erythrocyte and merozoite surfaces induces parasite inhibitory antibodies. J Exp Med. 2021 218(9):e20200170.